ISHII Jun

Engineering Biology Research CenterAssociate Professor
Faculty of Engineering / Department of Chemical Science and Engineering
Graduate School of Science, Technology and Innovation / Department of Science, Technology and Innovation

Research news

Research Keyword

  • Synthetic Biology
  • Metabolic Engineering
  • Genome Engineering
  • Gene Expression Control
  • Biosensor
  • Bioproduction
  • Microorganism
  • Biochemical Engineering

Research Areas

  • Manufacturing technology (mechanical, electrical/electronic, chemical engineering) / Applied biofunctional and bioprocess engineering

Committee History

  • Jun. 2021 - Present, 日本生物工学会 関西支部, 支部幹事(企画)
  • 2021 - Present, Frontiers in Chemical Engineering, Associate Editor
  • Jun. 2017 - May 2021, The Society for Biotechnology, Japan (SBJ), Editorial Board (Journal of Bioscience and Bioengineering)
  • Apr. 2017 - Mar. 2020, 文部科学省 科学技術・学術政策研究所 科学技術予測センター, 専門調査員

Award

  • Oct. 2023 神戸大学, 令和5年度学長表彰(財務貢献者)
    石井純

  • Jun. 2023 神戸大学大学院科学技術イノベーション研究科, 令和5年度研究科優秀教育賞
    石井純

  • Oct. 2022 神戸大学, 令和4年度学長表彰(財務貢献者)
    石井純

  • Oct. 2021 日本生物工学会, 第73回日本生物工学会大会トピックス賞, 光エネルギーを利用した大腸菌におけるメバロン酸のイソプレノールへの変換
    佐野海瑚人, 田中涼, 鎌田健太郎, 松田史生, 石井純, 弘埜陽子, 原清敬, 戸谷吉博, 清水浩
    Japan society

Paper

  • Christopher J. Vavricka, Masao Mochizuki, Satoshi Yuzawa, Masahiro Murata, Takanobu Yoshida, Naoki Watanabe, Masahiko Nakatsui, Jun Ishii, Kiyotaka Hara, Hal S. Alper, Tomohisa Hasunuma, Akihiko Kondo, Michihiro Araki
    Dec. 2024

  • Ririka Asama, Masahiro Tominaga, Sayaka Ito, Yoichiro Ito, Kazuhiro Takemura, Shun Sakuraba, Kohei Katsurada, Nobuo Fukuda, Akihiko Kondo, Jun Ishii
    Nov. 2024, Journal of Bioscience and Bioengineering
    Scientific journal

  • Ririka Asama, Cher J S Liu, Masahiro Tominaga, Yu-Ru Cheng, Yasuyuki Nakamura, Akihiko Kondo, Hsiang-Yu Wang, Jun Ishii
    BACKGROUND: Single-cell droplet microfluidics is an important platform for high-throughput analyses and screening because it provides an independent and compartmentalized microenvironment for reaction or cultivation by coencapsulating individual cells with various molecules in monodisperse microdroplets. In combination with microbial biosensors, this technology becomes a potent tool for the screening of mutant strains. In this study, we demonstrated that a genetically engineered yeast strain that can fluorescently sense agonist ligands via the heterologous expression of a human G-protein-coupled receptor (GPCR) and concurrently secrete candidate peptides is highly compatible with single-cell droplet microfluidic technology for the high-throughput screening of new agonistically active peptides. RESULTS: The water-in-oil microdroplets were generated using a flow-focusing microfluidic chip to encapsulate engineered yeast cells coexpressing a human GPCR [i.e., angiotensin II receptor type 1 (AGTR1)] and a secretory agonistic peptide [i.e., angiotensin II (Ang II)]. The single yeast cells cultured in the droplets were then observed under a microscope and analyzed using image processing incorporating machine learning techniques. The AGTR1-mediated signal transduction elicited by the self-secreted Ang II peptide was successfully detected via the expression of a fluorescent reporter in single-cell yeast droplet cultures. The system could also distinguish Ang II analog peptides with different agonistic activities. Notably, we further demonstrated that the microenvironment of the single-cell droplet culture enabled the detection of rarely existing positive (Ang II-secreting) yeast cells in the model mixed cell library, whereas the conventional batch-culture environment using a shake flask failed to do so. Thus, our approach provided compartmentalized microculture environments, which can prevent the diffusion, dilution, and cross-contamination of peptides secreted from individual single yeast cells for the easy identification of GPCR agonists. CONCLUSIONS: We established a droplet-based microfluidic platform that integrated an engineered yeast biosensor strain that concurrently expressed GPCR and self-secreted the agonistic peptides. This offers individually isolated microenvironments that allow the culture of single yeast cells secreting these peptides and gaging their signaling activities, for the high-throughput screening of agonistic peptides. Our platform base on yeast GPCR biosensors and droplet microfluidics will be widely applicable to metabolic engineering, environmental engineering, and drug discovery.
    Corresponding, Apr. 2024, Microbial cell factories, 23(1) (1), 104 - 104, English, International magazine, Co-authored internationally
    [Refereed]
    Scientific journal

  • Chinatsu Matsuyama, Taisuke Seike, Nobuyuki Okahashi, Teppei Niide, Kiyotaka Y. Hara, Yoko Hirono-Hara, Jun Ishii, Hiroshi Shimizu, Yoshihiro Toya, Fumio Matsuda
    Mar. 2024, Journal of Bioscience and Bioengineering
    [Refereed]
    Scientific journal

  • Kenta Morita, Musashi Takenaka, Kohei Tomita, Jun Ishii, Hideo Kawaguchi, Daisuke Murakami, Hikaru Amo, Miku Fujii, Tatsuo Maruyama, Takuya Matsumoto, Takashi Nishino, Chiaki Ogino
    Abstract Cellulose has been developed as an alternative to petrochemical materials. By comparison with refined nanofibers (RCNFs), lignocellulose nanofibers (LCNFs) show particular promise because it is produced from biomass using only mild pretreatment. The mechanical properties of LCNFs depend on the contained lignin. However, the microscopic location of the lignin contained in LCNFs has not been determined. Thus, we developed two methods to detect and visualize lignin. One uses a scanning transmission electron microscope (STEM) equipped with an energy dispersive X-ray spectroscopy detector. The other method uses an atomic force microscope (AFM) equipped with a cantilever coated with an aromatic molecule. Both methods revealed that the lignin in LCNFs covers a thin cellulose fiber and is precipitated in a grained structure. In particular, the AFM system was able to determine the nanoscopic location of lignin-rich areas. The present study establishes a strong tool for analyzing the characteristics of lignin-containing materials. Graphical abstract
    Springer Science and Business Media LLC, Nov. 2023, Cellulose, 30(18) (18), 11357 - 11367
    [Refereed]
    Scientific journal

  • Masahiko Sugimura, Taisuke Seike, Nobuyuki Okahashi, Yoshihiro Izumi, Takeshi Bamba, Jun Ishii, Fumio Matsuda
    Saccharomyces cerevisiae is a promising host for the bioproduction of higher alcohols, such as 2,3-butanediol (2,3-BDO). Metabolically engineered S. cerevisiae strains that produce 2,3-BDO via glycolysis have been constructed. However, the specific 2,3-BDO production rates of engineered strains must be improved. To identify approaches to improving the 2,3-BDO production rate, we investigated the factors contributing to higher ethanol production rates in certain industrial strains of S. cerevisiae compared to laboratory strains. Sequence analysis of 11 industrial strains revealed the accumulation of many nonsynonymous substitutions in RIM15, a negative regulator of high fermentation capability. Comparative metabolome analysis suggested a positive correlation between the rate of ethanol production and the activity of the pyruvate-consuming pathway. Based on these findings, RIM15 was deleted, and the pyruvate-consuming pathway was activated in YHI030, a metabolically engineered S. cerevisiae strain that produces 2,3-BDO. The titer, specific production rate, and yield of 2,3-BDO in the test tube-scale culture using the YMS106 strain reached 66.4 ± 4.4 mM, 1.17 ± 0.017 mmol (g dry cell weight h)−1, and 0.70 ± 0.03 mol (mol glucose consumed)−1. These values were 2.14-, 2.92-, and 1.81-fold higher than those of the vector control, respectively. These results suggest that bioalcohol production via glycolysis can be enhanced in a metabolically engineered S. cerevisiae strain by deleting RIM15 and activating the pyruvate-consuming pathway.
    MDPI AG, Nov. 2023, International Journal of Molecular Sciences, 24(22) (22), 16378 - 16378
    [Refereed]
    Scientific journal

  • Futa Yatabe, Taisuke Seike, Nobuyuki Okahashi, Jun Ishii, Fumio Matsuda
    Abstract Background “ATP wasting” has been observed in 13C metabolic flux analyses of Saccharomyces cerevisiae, a yeast strain commonly used to produce ethanol. Some strains of S. cerevisiae, such as the sake strain Kyokai 7, consume approximately two-fold as much ATP as laboratory strains. Increased ATP consumption may be linked to the production of ethanol, which helps regenerate ATP. Results This study was conducted to enhance ethanol and 2,3-butanediol (2,3-BDO) production in the S. cerevisiae strains, ethanol-producing strain BY318 and 2,3-BDO-producing strain YHI030, by expressing the fructose-1,6-bisphosphatase (FBPase) and ATP synthase (ATPase) genes to induce ATP dissipation. The introduction of a futile cycle for ATP consumption in the pathway was achieved by expressing various FBPase and ATPase genes from Escherichia coli and S. cerevisiae in the yeast strains. The production of ethanol and 2,3-BDO was evaluated using high-performance liquid chromatography and gas chromatography, and fermentation tests were performed on synthetic media under aerobic conditions in batch culture. The results showed that in the BY318-opt_ecoFBPase (expressing opt_ecoFBPase) and BY318-ATPase (expressing ATPase) strains, specific glucose consumption was increased by 30% and 42%, respectively, and the ethanol production rate was increased by 24% and 45%, respectively. In contrast, the YHI030-opt_ecoFBPase (expressing opt_ecoFBPase) and YHI030-ATPase (expressing ATPase) strains showed increased 2,3-BDO yields of 26% and 18%, respectively, and the specific production rate of 2,3-BDO was increased by 36%. Metabolomic analysis confirmed the introduction of the futile cycle. Conclusion ATP wasting may be an effective strategy for improving the fermentative biosynthetic capacity of S. cerevisiae, and increased ATP consumption may be a useful tool in some alcohol-producing strains.
    Springer Science and Business Media LLC, Oct. 2023, Microbial Cell Factories, 22(1) (1), International magazine
    [Refereed]
    Scientific journal

  • Toshiharu Arai, Mayumi Wada, Hiroki Nishiguchi, Yasushi Takimura, Jun Ishii
    Abstract Background The filamentous fungus Trichoderma reesei has been used as a host organism for the production of lignocellulosic biomass-degrading enzymes. Although this microorganism has high potential for protein production, it has not yet been widely used for heterologous recombinant protein production. Transcriptional induction of the cellulase genes is essential for high-level protein production in T. reesei; however, glucose represses this transcriptional induction. Therefore, cellulose is commonly used as a carbon source for providing its degraded sugars such as cellobiose, which act as inducers to activate the strong promoters of the major cellulase (cellobiohydrolase 1 and 2 (cbh1 and cbh2) genes. However, replacement of cbh1 and/or cbh2 with a gene encoding the protein of interest (POI) for high productivity and occupancy of recombinant proteins remarkably impairs the ability to release soluble inducers from cellulose, consequently reducing the production of POI. To overcome this challenge, we first used an inducer-free biomass-degrading enzyme expression system, previously developed to produce cellulases and hemicellulases using glucose as the sole carbon source, for recombinant protein production using T. reesei. Results We chose endogenous secretory enzymes and heterologous camelid small antibodies (nanobody) as model proteins. By using the inducer-free strain as a parent, replacement of cbh1 with genes encoding two intrinsic enzymes (aspartic protease and glucoamylase) and three different nanobodies (1ZVH, caplacizumab, and ozoralizumab) resulted in their high secretory productions using glucose medium without inducers such as cellulose. Based on signal sequences (carrier polypeptides) and protease inhibitors, additional replacement of cbh2 with the nanobody gene increased the percentage of POI to about 20% of total secreted proteins in T. reesei. This allowed the production of caplacizumab, a bivalent nanobody, to be increased to 9.49-fold (508 mg/L) compared to the initial inducer-free strain. Conclusions In general, whereas the replacement of major cellulase genes leads to extreme decrease in the degradation capacity of cellulose, our inducer-free system enabled it and achieved high secretory production of POI with increased occupancy in glucose medium. This system would be a novel platform for heterologous recombinant protein production in T. reesei.
    Springer Science and Business Media LLC, May 2023, Microbial Cell Factories, 22(1) (1), English
    [Refereed]
    Scientific journal

  • Kenta Morita, Yuya Nishimura, Jun Ishii, Tatsuo Maruyama
    The discovery of novel antibacterial drugs against infectious diseases has decreased over the past few decades because of their poor cost performance. In this study, we report that the nanoassembly of a short-peptide hydrogelator (P1) endowed novel antifungal selectivity to a conventional antifungal drug, amphotericin B (AmB), which expands its application spectrum. Clinical use of AmB is limited because of poor water solubility and poor selectivity in its toxicity, which often causes harmful effects on tissues. P1 was the low-molecular-weight hydrogelator (LMWHg) that showed low cytotoxicity and was enzymatically degraded. In general, an LMWHg entraps foreign hydrophobic molecules inside the hydrophobic space in the self-assembled body. P1 successfully solubilized AmB in water as a form of a nanocomplex (NC) that had a chain-like structure. The NCs showed remarkably low toxicity toward Saccharomyces cerevisiae as a model fungus when compared with free AmB, meaning that P1 suppressed the antifungal activity of AmB via coassembly. The suppressed antifungal activity of AmB recovered when P1 in the NCs was degraded by a protease to liberate AmB from the coassembly with P1. P1 at a high concentration formed a hydrogel incorporating AmB (AmB-P1 gel), in which the antifungal activity of AmB was suppressed as well as that in the NC. The coassembly with AmB affected the morphology of the P1 self-assembly. While S. cerevisiae that did not secrete proteases formed a colony on the AmB-P1 gel, Aspergillus oryzae that secreted proteases did not grow on the AmB-P1 gel at all, resulting in the selective killing of the fungus. Because some of malignant, infectious fungi secrete proteases, the coassembly strategy of conventional antifungal drugs with self-assembling molecules should lead to "drug repositioning" of approved drugs in the health and medical fields.
    American Chemical Society (ACS), Jan. 2023, ACS Applied Nano Materials, 6(2) (2), 1432 - 1440, English
    [Refereed]
    Scientific journal

  • Musashi Takekana, Takanobu Yoshida, Erika Yoshida, Sumika Ono, Shinnosuke Horie, Christopher J. Vavricka, Moe Hiratani, Kenji Tsuge, Jun Ishii, Yoshihiro Hayakawa, Akihiko Kondo, Tomohisa Hasunuma
    Elsevier BV, Jan. 2023, Journal of Chromatography B, 1215, 123588 - 123588, English
    [Refereed]
    Scientific journal

  • Mikoto Sano, Ryo Tanaka, Kentaro Kamata, Yoko Hirono-Hara, Jun Ishii, Fumio Matsuda, Kiyotaka Y. Hara, Hiroshi Shimizu, Yoshihiro Toya
    Bioconversion of key intermediate metabolites such as mevalonate into various useful chemicals is a promising strategy for microbial production. However, the conversion of mevalonate into isoprenoids requires a supply of adenosine triphosphate (ATP). Light-driven ATP regeneration using microbial rhodopsin is an attractive module for improving the intracellular ATP supply. In the present study, we demonstrated the ATP-consuming conversion of mevalonate to isoprenol using rhodopsin-expressing Escherichia coli cells as a whole-cell catalyst in a medium that does not contain energy cosubstrate, such as glucose. Heterologous genes for the synthesis of isoprenol from mevalonate, which requires three ATP molecules for the series of reactions, and a delta-rhodopsin gene derived from Haloterrigena turkmenica were cointroduced into E. coli. To evaluate the conversion efficiency of mevalonate to isoprenol, the cells were suspended in a synthetic medium containing mevalonate as the sole carbon source and incubated under dark or light illumination (100 μmol m-2 s-1). The specific isoprenol production rates were 10.0 ± 0.9 and 20.4 ± 0.7 μmol gDCW-1 h-1 for dark and light conditions, respectively. The conversion was successfully enhanced under the light condition. Furthermore, the conversion efficiency increased with increasing illumination intensity, suggesting that ATP regenerated by the proton motive force generated by rhodopsin using light energy can drive ATP-consuming reactions in the whole-cell catalyst.
    American Chemical Society (ACS), Dec. 2022, ACS Synthetic Biology, 11(12) (12), 3966 - 3972, English, International magazine
    [Refereed]
    Scientific journal

  • Kensuke Otsuka, Taisuke Seike, Yoshihiro Toya, Jun Ishii, Yoko Hirono-Hara, Kiyotaka Y Hara, Fumio Matsuda
    A light-driven ATP regeneration system using rhodopsin has been utilized as a method to improve the production of useful substances by microorganisms. To enable the industrial use of this system, the proton pumping rate of rhodopsin needs to be enhanced. Nonetheless, a method for this enhancement has not been established. In this study, we attempted to develop an evolutionary engineering method to improve the proton-pumping activity of rhodopsins. We first introduced random mutations into delta-rhodopsin (dR) from Haloterrigena turkmenica using error-prone PCR to generate approximately 7000 Escherichia coli strains carrying the mutant dR genes. Rhodopsin-expressing E. coli with enhanced proton pumping activity have significantly increased survival rates in prolonged saline water. Considering this, we enriched the mutant E. coli cells with higher proton pumping rates by selecting populations able to survive starvation under 50 μmol m-2 s-1 at 37 °C. As a result, we successfully identified two strains, in which proton pumping activity was enhanced two-fold by heterologous expression in E. coli in comparison to wild-type strains. The combined approach of survival testing using saline water and evolutionary engineering methods used in this study will contribute greatly to the discovery of a novel rhodopsin with improved proton pumping activity. This will facilitate the utilization of rhodopsin in industrial applications.
    Elsevier BV, Sep. 2022, Journal of bioscience and bioengineering, 134(6) (6), 484 - 490, English, Domestic magazine
    [Refereed]
    Scientific journal

  • Yoichiro Ito, Misa Ishigami, Noriko Hashiba, Yasuyuki Nakamura, Goro Terai, Tomohisa Hasunuma, Jun Ishii, Akihiko Kondo
    Corresponding, Wiley, Sep. 2022, Microbial Biotechnology, 15(9) (9), 2364 - 2378, English
    [Refereed]
    Scientific journal

  • Shota Isogai, Masahiro Tominaga, Akihiko Kondo, Jun Ishii
    Flavonoids, a major group of secondary metabolites in plants, are promising for use as pharmaceuticals and food supplements due to their health-promoting biological activities. Industrial flavonoid production primarily depends on isolation from plants or organic synthesis, but neither is a cost-effective or sustainable process. In contrast, recombinant microorganisms have significant potential for the cost-effective, sustainable, environmentally friendly, and selective industrial production of flavonoids, making this an attractive alternative to plant-based production or chemical synthesis. Structurally and functionally diverse flavonoids are derived from flavanones such as naringenin, pinocembrin and eriodictyol, the major basic skeletons for flavonoids, by various modifications. The establishment of flavanone-producing microorganisms can therefore be used as a platform for producing various flavonoids. This review summarizes metabolic engineering and synthetic biology strategies for the microbial production of flavanones. In addition, we describe directed evolution strategies based on recently-developed high-throughput screening technologies for the further improvement of flavanone production. We also describe recent progress in the microbial production of structurally and functionally complicated flavonoids via the flavanone modifications. Strategies based on synthetic biology will aid more sophisticated and controlled microbial production of various flavonoids.
    Frontiers Media SA, Jul. 2022, Frontiers in Chemical Engineering, 4, 880694, English
    [Refereed]
    Scientific journal

  • Masahiro Tominaga, Keita Miyazaki, Shoko Hataya, Yasumasa Mitsui, Shuji Kuroda, Akihiko Kondo, Jun Ishii
    Fermentative production of squalene in yeast as an alternative approach to extracting squalene from sharks or plants has attracted significant interest. However, squalene accumulation is limited due to its inevitable high-flux allocation toward ergosterol synthesis. In this study, we described expression control of squalene monooxygenase (Erg1p), the first-step enzyme of ergosterol synthesis from squalene, to significantly reduce squalene loss. We replaced the ERG1 promoter (PERG1) with three natural yeast promoters with different activities (PPCL2, PHCM1, and PTHI2). ERG1 controlled by PTHI2 showed 20 times higher squalene production compared with the wild-type strain, whereas the other two strains exhibited no significant difference. By combining the overexpression of rate-limiting enzyme and the deletion of non-essential competing pathway gene, the yeast Saccharomyces cerevisiae produced up to 379 mg/L of squalene.
    Elsevier BV, Jul. 2022, Journal of Bioscience and Bioengineering, 134(1) (1), 1 - 6, English, Domestic magazine
    [Refereed]
    Scientific journal

  • Yoshihiro Toya, Yoko Hirono-Hara, Hidenobu Hirayama, Kentaro Kamata, Ryo Tanaka, Mikoto Sano, Sayaka Kitamura, Kensuke Otsuka, Rei Abe-Yoshizumi, Satoshi P. Tsunoda, Hiroshi Kikukawa, Hideki Kandori, Hiroshi Shimizu, Fumio Matsuda, Jun Ishii, Kiyotaka Y. Hara
    In microbial fermentative production, ATP regeneration, while crucial for cellular processes, conflicts with efficient target chemical production because ATP regeneration exhausts essential carbon sources also required for target chemical biosynthesis. To wrestle with this dilemma, we harnessed the power of microbial rhodopsins with light-driven proton pumping activity to supplement with ATP, thereby facilitating the bioproduction of various chemicals. We first demonstrated a photo-driven ATP supply and redistribution of metabolic carbon flows to target chemical synthesis by installing already-known delta rhodopsin (dR) in Escherichia coli. In addition, we identified novel rhodopsins with higher proton pumping activities than dR, and created an engineered cell for in vivo self-supply of the rhodopsin-activator, all-trans-retinal. Our concept exploiting the light-powering ATP supplier offers a potential increase in carbon use efficiency for microbial productions through metabolic reprogramming.
    Elsevier, Jul. 2022, Metabolic Engineering, 72, 227 - 236, English, International magazine
    [Refereed]
    Scientific journal

  • Yoichiro Ito, Misa Ishigami, Goro Terai, Yasuyuki Nakamura, Noriko Hashiba, Teruyuki Nishi, Hikaru Nakazawa, Tomohisa Hasunuma, Kiyoshi Asai, Mitsuo Umetsu, Jun Ishii, Akihiko Kondo
    Abstract Expression of secreted recombinant proteins burdens the protein secretion machinery, limiting production. Here, we describe an approach to improving protein production by the non-conventional yeast Komagataella phaffii comprised of genome-wide screening for effective gene disruptions, combining them in a single strain, and recovering growth reduction by adaptive evolution. For the screen, we designed a multiwell-formatted, streamlined workflow to high-throughput assay of secretion of a single-chain small antibody, which is cumbersome to detect but serves as a good model of proteins that are difficult to secrete. Using the consolidated screening system, we evaluated >19,000 mutant strains from a mutant library prepared by a modified random gene-disruption method, and identified six factors for which disruption led to increased antibody production. We then combined the disruptions, up to quadruple gene knockouts, which appeared to contribute independently, in a single strain and observed an additive effect. Target protein and promoter were basically interchangeable for the effects of knockout genes screened. We finally used adaptive evolution to recover reduced cell growth by multiple gene knockouts and examine the possibility for further enhancing protein secretion. Our successful, three-part approach holds promise as a method for improving protein production by non-conventional microorganisms.
    Corresponding, Springer Science and Business Media LLC, Jun. 2022, Communications Biology, 5(1) (1), 561, English, International magazine
    [Refereed]
    Scientific journal

  • Masahiro Tominaga, Akihiko Kondo, Jun Ishii
    Transcriptional switches can be utilized for many purposes in synthetic biology, including the assembly of complex genetic circuits to achieve sophisticated cellular systems and the construction of biosensors for real-time monitoring of intracellular metabolite concentrations. Although to date such switches have mainly been developed in prokaryotes, those for eukaryotes are increasingly being reported as both rational and random engineering technologies mature. In this review, we describe yeast transcriptional switches with different modes of action and how to alter their properties. We also discuss directed evolution technologies for the rapid and robust construction of yeast transcriptional switches.
    Corresponding, {MDPI} {AG}, Apr. 2022, Life, 12(4) (4), 557 - 557, Japanese, International magazine
    [Refereed][Invited]
    Scientific journal

  • Christopher J. Vavricka, Shunsuke Takahashi, Naoki Watanabe, Musashi Takenaka, Mami Matsuda, Takanobu Yoshida, Ryo Suzuki, Hiromasa Kiyota, Jianyong Li, Hiromichi Minami, Jun Ishii, Kenji Tsuge, Michihiro Araki, Akihiko Kondo, Tomohisa Hasunuma
    Engineering the microbial production of secondary metabolites is limited by the known reactions of correctly annotated enzymes. Therefore, the machine learning discovery of specialized enzymes offers great potential to expand the range of biosynthesis pathways. Benzylisoquinoline alkaloid production is a model example of metabolic engineering with potential to revolutionize the paradigm of sustainable biomanufacturing. Existing bacterial studies utilize a norlaudanosoline pathway, whereas plants contain a more stable norcoclaurine pathway, which is exploited in yeast. However, committed aromatic precursors are still produced using microbial enzymes that remain elusive in plants, and additional downstream missing links remain hidden within highly duplicated plant gene families. In the current study, machine learning is applied to predict and select plant missing link enzymes from homologous candidate sequences. Metabolomics-based characterization of the selected sequences reveals potential aromatic acetaldehyde synthases and phenylpyruvate decarboxylases in reconstructed plant gene-only benzylisoquinoline alkaloid pathways from tyrosine. Synergistic application of the aryl acetaldehyde producing enzymes results in enhanced benzylisoquinoline alkaloid production through hybrid norcoclaurine and norlaudanosoline pathways.
    Springer Nature, Mar. 2022, Nature Communications, 13(1) (1), 1405 - 1405, English, International magazine, Co-authored internationally
    [Refereed]
    Scientific journal

  • Keisuke Morita, Taisuke Seike, Jun Ishii, Fumio Matsuda, Hiroshi Shimizu
    Saccharomyces cerevisiae has been widely used in bioproduction. To produce a target product other than ethanol, ethanol production must be decreased to enhance target production. An ethanol non-producing yeast strain was previously constructed by knocking out pyruvate decarboxylase (PDC) genes in the ethanol synthetic pathway. However, glucose uptake by the ethanol-non-producing yeast strain was significantly decreased. In this study, dead Cas9 (dCas9) was used to reduce ethanol synthesis during 2,3-butanediol production without reduction of glucose. The binding site of guide RNA used to effectively suppress PDC1 promoter-driven red fluorescent protein expression by dCas9 was identified and applied to control PDC1 expression. The production of 2,3-butanediol rather than ethanol was improved in repetitive test tube culture. Additionally, ethanol production was decreased and 2,3-butanediol production was increased in the strain expressing dCas9 targeting the PDC1 promoter in the third round of cultivation, compared with the control strain.
    Elsevier, Mar. 2022, Journal of Bioscience and Bioengineering, 133(3) (3), 208 - 212, English, International magazine
    [Refereed]
    Scientific journal

  • Teruyuki Nishi, Yoichiro Ito, Yasuyuki Nakamura, Taiki Yamaji, Noriko Hashiba, Masaya Tamai, Yoshihiko Yasohara, Jun Ishii, Akihiko Kondo
    The methylotrophic yeast species Komagataella phaffii (synonym: Pichia pastoris) is widely used as a host for recombinant protein production. Although several genetic engineering techniques are being employed on K. phaffii, advanced methods such as in vivo DNA assembly in this yeast species are required for synthetic biology applications. In this study, we established a technique for accomplishing one-step in vivo assembly of multiple DNA fragments and genomic integration in K. phaffii. To concurrently achieve an accurate multiple DNA assembly and a high-efficient integration into the target genomic locus in vivo, a K. phaffii strain, lacking a non-homologous end joining-related protein, DNA ligase IV (Dnl4p), that has been reported to improve gene targeting efficiency by homologous recombination, was used. Using green fluorescent protein along with the lycopene biosynthesis, we showed that our method that included a Dnl4p-defective strain permits direct and easy engineering of K. phaffii strains.
    Corresponding, American Chemical Society ({ACS}), Jan. 2022, ACS Synthetic Biology, 11(2) (2), 644 - 654, English, International magazine
    [Refereed]
    Scientific journal

  • Kohei Katsurada, Masahiro Tominaga, Misato Kaishima, Hiroko Kato, Toshihide Matsuno, Chiaki Ogino, Akihiko Kondo, Jun Ishii, Katsumi Takayama
    We describe a novel expression cassette that enables efficient and constitutive expression of the ZZ domain derived from Staphylococcus aureus protein A on the yeast cell surface to easily prepare yeast-based immunosorbents. Using this expression cassette containing the PGK1 promoter, a secretion signal derived from α-factor, and a Flo1-derived anchor protein, we successfully created a yeast-based immunosorbent for human serum albumin.
    Corresponding, Microbiology Research Foundation, Dec. 2021, The Journal of General and Applied Microbiology, 67(6) (6), 265 - 268, English, International magazine
    [Refereed]
    Scientific journal

  • Erika Yoshida, Motoki Kojima, Munenori Suzuki, Fumio Matsuda, Kazutaka Shimbo, Akiko Onuki, Yousuke Nishio, Yoshihiro Usuda, Akihiko Kondo, Jun Ishii
    (-)-Carvone is a monoterpenoid with a spearmint flavor. A sustainable biotechnological production process for (-)-carvone is desirable. Although all enzymes in (-)-carvone biosynthesis have been functionally expressed in Escherichia coli independently, the yield was low in previous studies. When cytochrome P450 limonene-6-hydroxylase (P450)/cytochrome P450 reductase (CPR) and carveol dehydrogenase (CDH) were expressed in a single strain, by-product formation (dihydrocarveol and dihydrocarvone) was detected. We hypothesized that P450 and CDH expression levels differ in E. coli. Thus, two strains independently expressing P450/CPR and CDH were mixed with different ratios, confirming increased carvone production and decreased by-product formation when CDH input was reduced. The optimum ratio of enzyme expression to maximize (-)-carvone production was determined using the proteome analysis quantification concatamer (QconCAT) method. Thereafter, a single strain expressing both P450/CPR and CDH was constructed to imitate the optimum expression ratio. The upgraded strain showed a 15-fold improvement compared to the initial strain, showing a 44 ± 6.3 mg/L (-)-carvone production from 100 mg/L (-)-limonene. Our study showed the usefulness of the QconCAT proteome analysis method for strain development in the industrial biotechnology field.
    Spring Nature, Nov. 2021, Scientific reports, 11(1) (1), 22126 - 22126, English, International magazine
    [Refereed]
    Scientific journal

  • Shota Isogai, Nobuyuki Okahashi, Ririka Asama, Tomomi Nakamura, Tomohisa Hasunuma, Fumio Matsuda, Jun Ishii, Akihiko Kondo
    Reconstitution of prenylflavonoids using the flavonoid biosynthetic pathway and prenyltransferases (PTs) in microbes can be a promising attractive alternative to plant-based production or chemical synthesis. Here, we demonstrate that promiscuous microbial PTs can be a substitute for regiospecific but mostly unidentified botanical PTs. To test the prenylations of naringenin, we constructed a yeast strain capable of producing naringenin from l-phenylalanine by genomic integration of six exogenous genes encoding components of the naringenin biosynthetic pathway. Using this platform strain, various microbial PTs were tested for prenylnaringenin production. In vitro screening demonstrated that the fungal AnaPT (a member of the tryptophan dimethylallyltransferase family) specifically catalyzed C-3' prenylation of naringenin, whereas SfN8DT-1, a botanical PT, specifically catalyzed C-8 prenylation. In vivo, the naringenin-producing strain expressing the microbial AnaPT exhibited heterologous microbial production of 3'-prenylnaringenin (3'-PN), in contrast to the previously reported in vivo production of 8-prenylnaringenin (8-PN) using the botanical SfN8DT-1. These findings provide strategies towards expanding the production of a variety of prenylated compounds, including well-known prenylnaringenins and novel prenylflavonoids. These results also suggest the opportunity for substituting botanical PTs, both known and unidentified, that display relatively strict regiospecificity of the prenyl group transfer.
    Corresponding, Elsevier, Jun. 2021, Metabolic Engineering Communications, 12, e00169 - e00169, English, International magazine
    [Refereed]
    Scientific journal

  • Shinichiro Shoji, Taiki Yamaji, Harumi Makino, Jun Ishii, Akihiko Kondo
    β-Nicotinamide mononucleotide (NMN) is, one of the nucleotide compounds, a precursor of NAD+ and has recently attracted attention as a nutraceutical. Here, we develop a whole-cell biocatalyst using Escherichia coli, which enabled selective and effective high production of NMN from the inexpensive feedstock substrates glucose and nicotinamide (Nam). Notably, we identify two actively functional transporters (NiaP and PnuC) and a high-activity key enzyme (Nampt), permitting intracellular Nam uptake, efficient conversion of phosphoribosyl pyrophosphate (PRPP; supplied from glucose) and Nam to NMN, and NMN excretion extracellularly. Further, enhancement of the PRPP biosynthetic pathway and optimization of individual gene expression enable drastically higher NMN production than reported thus far. The strain extracellularly produces 6.79 g l−1 of NMN from glucose and Nam, and the reaction selectivity from Nam to NMN is 86%. Our approach will be promising for low-cost, high-quality industrial production of NMN and other nucleotide compounds using microorganisms.
    Elsevier {BV}, May 2021, Metabolic Engineering, 65, 167 - 177, English, International magazine
    [Refereed]
    Scientific journal

  • Masahiro Tominaga, Kenta Nozaki, Daisuke Umeno, Jun Ishii, Akihiko Kondo
    A wide repertoire of genetic switches has accelerated prokaryotic synthetic biology, while eukaryotic synthetic biology has lagged in the model organism Saccharomyces cerevisiae. Eukaryotic genetic switches are larger and more complex than prokaryotic ones, complicating the rational design and evolution of them. Here, we present a robust workflow for the creation and evolution of yeast genetic switches. The selector system was designed so that both ON- and OFF-state selection of genetic switches is completed solely by liquid handling, and it enabled parallel screen/selection of different motifs with different selection conditions. Because selection threshold of both ON- and OFF-state selection can be flexibly tuned, the desired selection conditions can be rapidly pinned down for individual directed evolution experiments without a prior knowledge either on the library population. The system’s utility was demonstrated using 20 independent directed evolution experiments, yielding genetic switches with elevated inducer sensitivities, inverted switching behaviours, sensory functions, and improved signal-to-noise ratio (>100-fold induction). The resulting yeast genetic switches were readily integrated, in a plug-and-play manner, into an AND-gated carotenoid biosynthesis pathway.
    Corresponding, Springer Nature, Mar. 2021, Nature Communications, 12(1) (1), 1846, English, International magazine
    [Refereed]
    Scientific journal

  • Yasuyuki Nakamura, Ririka Asama, Takuya Tabata, Kenta Morita, Tatsuo Maruyama, Akihiko Kondo, Jun Ishii
    Melatonin is an indoleamine neurohormone made by the pineal gland. Its receptors, MTNR1A and MTNR1B, are members of the G-protein-coupled receptor (GPCR) family and are involved in sleep, circadian rhythm, and mood disorders, and in the inhibition of cancer growth. These receptors, therefore, represent significant molecular targets for insomnia, circadian sleep disorders, and cancer. The yeast Saccharomyces cerevisiae is an attractive host for assaying agonistic activity for human GPCR. We previously constructed a GPCR-based biosensor employing a high-sensitivity yeast strain that incorporated both a chimeric yeast-human G alpha protein and a bright fluorescent reporter gene (ZsGreen). Similar approaches have been used for simple and convenient measurements of various GPCR activities. In the current study, we constructed a fluorescence-based yeast biosensor for monitoring the signaling activation of human melatonin receptors. We used this system to analyze point mutations, including previously unreported mutations of the consensus sequences of MTNR1A and MTNR1B melatonin receptors and compared their effects. Most mutations in the consensus sequences significantly affected the signaling capacities of both receptors, but several mutations showed differences between these subtype receptors. Thus, this yeast biosensor holds promise for revealing the functions of melatonin receptors.
    Wiley, Feb. 2021, Biotechnology and Bioengineering, 118(2) (2), 863 - 876, English, International magazine
    [Refereed]
    Scientific journal

  • Yoichiro Ito, Goro Terai, Misa Ishigami, Noriko Hashiba, Yasuyuki Nakamura, Takahiro Bamba, Ryota Kumokita, Tomohisa Hasunuma, Kiyoshi Asai, Jun Ishii, Akihiko Kondo
    Abstract In the yeast Saccharomyces cerevisiae, terminator sequences not only terminate transcription but also affect expression levels of the protein-encoded upstream of the terminator. The non-conventional yeast Pichia pastoris (syn. Komagataella phaffii) has frequently been used as a platform for metabolic engineering but knowledge regarding P. pastoris terminators is limited. To explore terminator sequences available to tune protein expression levels in P. pastoris, we created a ‘terminator catalog’ by testing 72 sequences, including terminators from S. cerevisiae or P. pastoris and synthetic terminators. Altogether, we found that the terminators have a tunable range of 17-fold. We also found that S. cerevisiae terminator sequences maintain function when transferred to P. pastoris. Successful tuning of protein expression levels was shown not only for the reporter gene used to define the catalog but also using betaxanthin production as an example application in pathway flux regulation. Moreover, we found experimental evidence that protein expression levels result from mRNA abundance and in silico evidence that levels reflect the stability of mRNA 3′-UTR secondary structure. In combination with promoter selection, the novel terminator catalog constitutes a basic toolbox for tuning protein expression levels in metabolic engineering and synthetic biology in P. pastoris.
    Corresponding, Oxford University Press ({OUP}), Dec. 2020, Nucleic Acids Research, 48(22) (22), 13000 - 13012, English, International magazine
    [Refereed]
    Scientific journal

  • Yuya Nishimura, Ryosuke Ezawa, Kenta Morita, Masao Nakayama, Jun Ishii, Ryohei Sasaki, Chiaki Ogino, Akihiko Kondo
    Radiosensitizing therapy for cancer treatment that enhances the effect of existing radiation therapy and enables noninvasive therapy has attracted attention. In this study, to achieve target cell-specific noninvasive cancer treatment using a Z(HER2)-bionanocapsule/liposome (BNC/LP), a carrier that binds to human epidermal growth factor receptor 2 (HER2), we evaluated the delivery of anticancer drugs and radiosensitizers and treatment effects in vitro and in vivo in mice. Target cell-specific cytotoxic activity and antitumor effects were confirmed following delivery of doxorubicin-encapsulated particles. In addition, cell damage due to radiosensitizing effects was confirmed in combination with X-ray irradiation following delivery of particles containing polyacrylic acid-modified titanium peroxide nanoparticles as a radiosensitizer. Furthermore, even when the particles were injected via the tail vein of mice, they accumulated in the tumor and exhibited an antitumor effect because of radiosensitization. Therefore, Z(HER2)-BNC/ LP is expected to be a carrier that releases small-molecule drugs into the target cell cytoplasm and delivers a radiosensitizer such as inorganic nanoparticles, enabling combination therapy with X-rays to the target tumor.
    American Chemical Society ({ACS}), Nov. 2020, ACS Applied Bio Materials, 3(11) (11), 7743 - 7751, English, International magazine
    [Refereed]
    Scientific journal

  • Keisuke Morita, Fumio Matsuda, Koji Okamoto, Jun Ishii, Akihiko Kondo, Hiroshi Shimizu
    BACKGROUND: Saccharomyces cerevisiae is a suitable host for the industrial production of pyruvate-derived chemicals such as ethanol and 2,3-butanediol (23BD). For the improvement of the productivity of these chemicals, it is essential to suppress the unnecessary pyruvate consumption in S. cerevisiae to redirect the metabolic flux toward the target chemical production. In this study, mitochondrial pyruvate transporter gene (MPC1) or the essential gene for mitophagy (ATG32) was knocked-out to repress the mitochondrial metabolism and improve the production of pyruvate-derived chemical in S. cerevisiae. RESULTS: The growth rates of both aforementioned strains were 1.6-fold higher than that of the control strain. 13C-metabolic flux analysis revealed that both strains presented similar flux distributions and successfully decreased the tricarboxylic acid cycle fluxes by 50% compared to the control strain. Nevertheless, the intracellular metabolite pool sizes were completely different, suggesting distinct metabolic effects of gene knockouts in both strains. This difference was also observed in the test-tube culture for 23BD production. Knockout of ATG32 revealed a 23.6-fold increase in 23BD titer (557.0 ± 20.6 mg/L) compared to the control strain (23.5 ± 12.8 mg/L), whereas the knockout of MPC1 revealed only 14.3-fold increase (336.4 ± 113.5 mg/L). Further investigation using the anaerobic high-density fermentation test revealed that the MPC1 knockout was more effective for ethanol production than the 23BD production. CONCLUSION: These results suggest that the engineering of the mitochondrial transporters and membrane dynamics were effective in controlling the mitochondrial metabolism to improve the productivities of chemicals in yeast cytosol.
    Oct. 2019, Microbial cell factories, 18(1) (1), 177 - 177, English, International magazine
    [Refereed]
    Scientific journal

  • Christopher J. Vavricka, Takanobu Yoshida, Yuki Kuriya, Shunsuke Takahashi, Teppei Ogawa, Fumie Ono, Kazuko Agari, Hiromasa Kiyota, Jianyong Li, Jun Ishii, Kenji Tsuge, Hiromichi Minami, Michihiro Araki, Tomohisa Hasunuma, Akihiko Kondo
    In the original version of this Article, the abbreviation of 3,4-dihydroxyphenylacetaldehyde synthase presented in the first paragraph of the Discussion section was given incorrectly as DYPAA. The correct abbreviation for this enzyme is DHPAAS. This error has been corrected in both the PDF and HTML versions of the Article.
    Springer Science and Business Media {LLC}, May 2019, Nature Communications, 10(1) (1), 2336 - 2336, English, International magazine
    [Refereed]
    Scientific journal

  • Christopher J. Vavricka, Takanobu Yoshida, Yuki Kuriya, Shunsuke Takahashi, Teppei Ogawa, Fumie Ono, Kazuko Agari, Hiromasa Kiyota, Jianyong Li, Jun Ishii, Kenji Tsuge, Hiromichi Minami, Michihiro Araki, Tomohisa Hasunuma, Akihiko Kondo
    © 2019, The Author(s). Previous studies have utilized monoamine oxidase (MAO) and L-3,4-dihydroxyphenylalanine decarboxylase (DDC) for microbe-based production of tetrahydropapaveroline (THP), a benzylisoquinoline alkaloid (BIA) precursor to opioid analgesics. In the current study, a phylogenetically distinct Bombyx mori 3,4-dihydroxyphenylacetaldehyde synthase (DHPAAS) is identified to bypass MAO and DDC for direct production of 3,4-dihydroxyphenylacetaldehyde (DHPAA) from L-3,4-dihydroxyphenylalanine (L-DOPA). Structure-based enzyme engineering of DHPAAS results in bifunctional switching between aldehyde synthase and decarboxylase activities. Output of dopamine and DHPAA products is fine-tuned by engineered DHPAAS variants with Phe79Tyr, Tyr80Phe and Asn192His catalytic substitutions. Balance of dopamine and DHPAA products enables improved THP biosynthesis via a symmetrical pathway in Escherichia coli. Rationally engineered insect DHPAAS produces (R,S)-THP in a single enzyme system directly from L-DOPA both in vitro and in vivo, at higher yields than that of the wild-type enzyme. However, DHPAAS-mediated downstream BIA production requires further improvement.
    Springer Science and Business Media {LLC}, May 2019, Nature Communications, 10(1) (1)
    [Refereed]
    Scientific journal

  • Koichi Tamano, Robert Sidney Cox, 3rd, Kenji Tsuge, Ai Miura, Ayano Itoh, Jun Ishii, Tomohiro Tamura, Akihiko Kondo, Masayuki Machida
    Free dihomo-γ-linolenic acid (DGLA) and its desaturated form, free arachidonic acid (ARA) are polyunsaturated free fatty acids (FFAs). They are useful raw materials to produce eicosanoid pharmaceuticals. In this study, we aimed at their production by the oleaginous filamentous fungus Aspergillus oryzae via metabolic engineering. Three genes encoding enzymes involved in the synthesis of DGLA and ARA, were isolated from the filamentous fungus Mortierella alpina that produces ARA in a triacylglycerol form. These genes were concatenated to promoters and terminators of highly expressed genes of A. oryzae, and the concatenated DNA fragments were further concatenated with each other to generate a single DNA fragment in the form of a biosynthetic gene cluster. By homologous recombination, the resulting DNA fragment was integrated to the chromosome of the A. oryzae acyl-CoA synthetase gene disruptant whose FFA productivity was enhanced at 9.2-fold more than the wild-type strain. The DNA-integrated disruptant produced free DGLA but did not produce free ARA. Thus, focusing on free DGLA, after removal of the gene for converting DGLA to ARA, the constructed strain produced free DGLA at 145 mg/l for 5 d. Also, by supplementing Triton X-100 surfactant at 1% to the culture, over 80% of free DGLA was released from cells without inhibiting the growth. Consequently, the constructed strain will be useful for attempting production of free DGLA-derived eicosanoids because it bypasses excision of free DGLA from triacylglycerols by lipase. To our knowledge, this is the first report on microbial production of free DGLA and its extracellular release.
    Elsevier {BV}, Apr. 2019, Journal of Bioscience and Bioengineering, 127(4) (4), 451 - 457, English, Domestic magazine
    [Refereed]
    Scientific journal

  • Hiroya Itoh, Makoto Matsui, Yuki Miyamura, Itaru Takeda, Jun Ishii, Toshitaka Kumagai, Masayuki Machida, Takashi Shibata, Masanori Arita
    American Chemical Society ({ACS}), Dec. 2018, ACS Synthetic Biology, 7(12) (12), 2783 - 2789, English, International magazine
    [Refereed]
    Scientific journal

  • Yoichiro Ito, Toru Watanabe, Shimpei Aikawa, Teruyuki Nishi, Tozo Nishiyama, Yasuyuki Nakamura, Tomohisa Hasunuma, Yuji Okubo, Jun Ishii, Akihiko Kondo
    The non-conventional yeast Komagataella phaffii, formerly Pichia pastoris, is a popular host for recombinant protein production. The relatively lower gene targeting efficiency observed in this species occurs due to high levels of non-homologous recombination activity. In the current study, we explored the function of the K. phaffii homolog of DNA ligase IV (Dnl4p) by creating a DNL4-disrupted strain. To assess the roles of non-homologous end joining (NHEJ)-related proteins in this species, strains deleted for either or both genes encoding Dnl4p or the telomeric Ku complex subunit (Ku70p) were generated. These deletions were constructed by either of two distinct marker-recycling methods (yielding either a seamless gene deletion or a Cre-loxP-mediated gene deletion). The resulting dnl4- and/or ku70-deleted K. phaffii strains were used to evaluate gene targeting efficiency in gene knock-out and gene knock-in experiments. The Dnl4p-defective strain showed improved gene targeting efficiency for homologous recombination compared to the wild-type and Ku70p-deffective strains. The dnl4 ku70 double knock-out strain exhibited a further improvement in gene targeting efficiency. Thus, the K. phaffii dnl4 and dnl4 ku70 deletion strains are expected to serve as useful platforms for functional analysis and strain development in this species.
    Corresponding, Oxford University Press ({OUP}), Nov. 2018, FEMS Yeast Research, 18(7) (7), foy074, English, International magazine
    [Refereed]
    Scientific journal

  • Yasuyuki Nakamura, Teruyuki Nishi, Risa Noguchi, Yoichiro Ito, Toru Watanabe, Tozo Nishiyama, Shimpei Aikawa, Tomohisa Hasunuma, Jun Ishii, Yuji Okubo, Akihiko Kondo, Frank E. Löffler
    The methylotrophic yeast Pichia pastoris is widely used to produce recombinant proteins, taking advantage of this species' high-density cell growth and strong ability to secrete proteins. Circular plasmids containing the P. pastoris-specific autonomously replicating sequence (PARS1) permit transformation of P. pastoris with higher efficiency than obtained following chromosomal integration by linearized DNA. Unfortunately, however, existing autonomously replicating plasmids are known to be inherently unstable. In this study, we used transcriptome sequencing (RNA-seq) data and genome sequence information to independently identify, on each of the four chromosomes, centromeric DNA sequences consisting of long inverted repeat sequences. By examining the chromosome 2 centromeric DNA sequence (Cen2) in detail, we demonstrate that an ∼111-bp region located at one end of the putative centromeric sequence had autonomous replication activity. In addition, the full-length Cen2 sequence, which contains two long inverted repeat sequences and a nonrepetitive central core region, is needed for the accurate replication and distribution of plasmids in P. pastoris. Thus, we constructed a new, stable, autonomously replicating plasmid vector that harbors the entire Cen2 sequence; this episome facilitates genetic manipulation in P. pastoris, providing high transformation efficiency and plasmid stability. IMPORTANCE Secretory production of recombinant proteins is the most important application of the methylotrophic yeast Pichia pastoris, a species that permits mass production of heterologous proteins. To date, the genetic engineering of P. pastoris has relied largely on integrative vectors due to the lack of user-friendly tools. Autonomously replicating Pichia plasmids are expected to facilitate genetic manipulation; however, the existing systems, which use autonomously replicating sequences (ARSs) such as the P. pastoris-specific ARS (PARS1), are known to be inherently unstable for plasmid replication and distribution. Recently, the centromeric DNA sequences of P. pastoris were identified in back-to-back studies published by several groups; therefore, a new episomal plasmid vector with centromere DNA as a tool for genetic manipulation of P. pastoris is ready to be developed.
    Corresponding, American Society for Microbiology, Aug. 2018, Applied and Environmental Microbiology, 84(15) (15), e02882 - 17, English, International magazine
    [Refereed]
    Scientific journal

  • Ishii, J., Morita, K., Ida, K., Kato, H., Kinoshita, S., Hataya, S., Shimizu, H., Kondo, A., Matsuda, F.
    Corresponding, BioMed Central, Jun. 2018, Biotechnology for Biofuels, 11(1) (1), 180, English, International magazine
    [Refereed]
    Scientific journal

  • Restu WK, Nishida Y, Yamamoto S, Ishii J, Maruyama T
    Jun. 2018, Langmuir : the ACS journal of surfaces and colloids, 34(27) (27), 8065 - 8074, English
    [Refereed]
    Scientific journal

  • Hiroki Hashi, Yasuyuki Nakamura, Jun Ishii, Akihiko Kondo
    Neurotensin receptor type 1 (NTSR1), a member of the G-protein-coupled receptor (GPCR) family, is naturally activated by binding of a neurotensin peptide, leading to a variety of physiological effects. The budding yeast Saccharomyces cerevisiae is a proven host organism for assaying the agonistic activation of human GPCRs. Previous studies showed that yeast cells can functionally express human NTSR1 receptor, permitting the detection of neurotensin-promoted signaling using a ZsGreen fluorescent reporter gene. However, the fluorescence intensity (sensitivity) of NTSR1-expressing yeast cells is low compared to that of yeast cells expressing other human GPCRs (e.g., human somatostatin receptors). The present study sought to increase the sensitivity of the NTSR1-expressing yeast for use as a fluorescent biosensor, including modification of the expression of human NTSR1 in yeast. Changes in the transcription, translation, and transport of the receptor are attempted by altering the promoter, consensus Kozak-like sequence, and secretion signal sequences of the NTSR1-encoding gene. The resulting yeast cells exhibited increased sensitivity to exogenously added peptide. The cells are further engineered by using cell-surface display technology to ensure that the agonistic peptides are secreted and tethered to the yeast cell wall, yielding cells with enhanced NTSR1 activation. This yeast biosensor holds promise for the identification of agonists to treat NTSR1-related diseases.
    Corresponding, Wiley-Blackwell, Apr. 2018, Biotechnology Journal, 13(4) (4), 1700522 - 1700522, English, International magazine
    [Refereed]
    Scientific journal

  • Nishimura, Y., Matsui, T., Ishii, J., Kondo, A.
    Background: To produce 1-propanol as a potential biofuel, metabolic engineering of microorganisms, such as E. coli, has been studied. However, 1-propanol production using metabolically engineered Saccharomyces cerevisiae, which has an amazing ability to produce ethanol and is thus alcohol-tolerant, has infrequently been reported. Therefore, in this study, we aimed to engineer S. cerevisiae strains capable of producing 1-propanol at high levels. Results: We found that the activity of endogenous 2-keto acid decarboxylase and alcohol/aldehyde dehydrogenase is sufficient to convert 2-ketobutyrate (2 KB) to 500 mg/L 1-propanol in yeast. Production of 1-propanol could be increased by: (i) the construction of an artificial 2 KB biosynthetic pathway from pyruvate via citramalate (cimA) (ii) overexpression of threonine dehydratase (tdcB) (iii) enhancement of threonine biosynthesis from aspartate (thrA, thrB and thrC) and (iv) deletion of the GLY1 gene that regulates a competing pathway converting threonine to glycine. With high-density anaerobic fermentation of the engineered S. cerevisiae strain YG5C4231, we succeeded in producing 180 mg/L 1-propanol from glucose. Conclusion: These results indicate that the engineering of a citramalate-mediated pathway as a production method for 1-propanol in S. cerevisiae is effective. Although optimization of the carbon flux in S. cerevisiae is necessary to harness this pathway, it is a promising candidate for the large-scale production of 1-propanol.
    BioMed Central Ltd., Mar. 2018, Microbial Cell Factories, 17(1) (1), 38, English, International magazine
    [Refereed]
    Scientific journal

  • Nambu-Nishida, Y., Sakihama, Y., Ishii, J., Hasunuma, T., Kondo, A.
    To efficiently utilize xylose, a major sugar component of hemicelluloses, in Saccharomyces cerevisiae requires the proper expression of varied exogenous and endogenous genes. To expand the repertoire of promoters in engineered xylose-utilizing yeast strains, we selected promoters in S. cerevisiae during cultivation and fermentation using xylose as a carbon source. To select candidate promoters that function in the presence of xylose, we performed comprehensive gene expression analyses using xylose-utilizing yeast strains both during xylose and glucose fermentation. Based on microarray data, we chose 29 genes that showed strong, moderate, and weak expression in xylose rather than glucose fermentation. The activities of these promoters in a xylose-utilizing yeast strain were measured by lacZ reporter gene assays over time during aerobic cultivation and microaerobic fermentation, both in xylose and glucose media. In xylose media, PTDH3, PFBA1, and PTDH1 were favorable for high expression, and PSED1, PHXT7, PPDC1, PTEF1, PTPI1, and PPGK1 were acceptable for medium–high expression in aerobic cultivation, and moderate expression in microaerobic fermentation. PTEF2 allowed moderate expression in aerobic culture and weak expression in microaerobic fermentation, although it showed medium–high expression in glucose media. PZWF1 and PSOL4 allowed moderate expression in aerobic cultivation, while showing weak but clear expression in microaerobic fermentation. PALD3 and PTKL2 showed moderate promoter activity in aerobic cultivation, but showed almost no activity in microaerobic fermentation. The knowledge of promoter activities in xylose cultivation obtained in this study will permit the control of gene expression in engineered xylose-utilizing yeast strains that are used for hemicellulose fermentation.
    Elsevier B.V., Jan. 2018, Journal of Bioscience and Bioengineering, 125(1) (1), 76 - 86, English, International magazine
    [Refereed]
    Scientific journal

  • Nobuo Fukuda, Misato Kaishima, Jun Ishii, Shinya Honda
    The yeast Saccharomyces cerevisiae is a useful eukaryotic host organism for studying GPCRs as monomolecular models. Fluorescent reporter gene assays for GPCRs provide a convenient assay for measuring receptor activity using fluorometric instruments. Generally, these assays detect receptor activation by agonistic ligands as the induction of fluorescent reporter expression, whereas antagonistic activities are detected by competition with agonistic ligands, resulting in decreases in fluorescence intensity. In the current study, we established a system for inverted expression of a fluorescent reporter by incorporating a PEST-tag and finding out a promoter inhibited by activation of the GPCR signaling pathway from yeast endogenous promoters. Because agonists prevent fluorescent reporter expression in this system, antagonists compete with agonists and yield increased fluorescence intensity. We used the yeast endogenous pheromone receptor as a model GPCR to demonstrate the feasibility of our system for positive detection targeted at antagonists. Compared to results when only agonists were added to yeast cells, more than 10-fold higher fluorescence intensity was observed when antagonists were added in combination with agonists. The approach described here has the potential to markedly accelerate the identification of GPCR antagonists by providing rapid and straightforward responses.
    AMER CHEMICAL SOC, Aug. 2017, ACS SYNTHETIC BIOLOGY, 6(8) (8), 1554 - 1562, English
    [Refereed]
    Scientific journal

  • Shintaro Ryo, Jun Ishii, Toshihide Matsuno, Yasuyuki Nakamura, Daiki Matsubara, Masahiro Tominaga, Akihiko Kondo
    The GAL expression system is the most frequently used induction technique in the yeast Saccharomyces cerevisiae. Here we report a simple but powerful genetic circuit for use with the GAL induction system. Briefly, an artificial positive feedback circuit was incorporated into the GAL regulatory network. We selected green fluorescent protein (GFP) as a reporter of GAL] induction, and designed a strain that expressed a constitutively active Gal3 mutant protein (Gal3(c)) under control of the GAL10 promoter. In the resulting strain, GALL and GAL10 promoters regulate the expression of GFP and GALS(c), respectively. Because Gal3(c) sequesters the Gal80 repressor away from the Gal4 transcriptional activator in the same manner as the galactose-bound Gal3, the expressed Gal3(c) protein provokes further expression of GFP and Gal3(c), yielding further enhancement of GAL induction. Thus, this GAL3(c)-mediated positive feedback circuit permits substantially enriched induction of a target gene at extremely low concentrations, or even in the absence, of galactose, while maintaining the strict glucose-mediated repression of the target.
    American Chemical Society ({ACS}), Jun. 2017, ACS Synthetic Biology, 6(6) (6), 928 - 935, English
    [Refereed]
    Scientific journal

  • Inokuma, K., Ishii, J., Hara, K.Y., Mochizuki, M., Hasunuma, T., Kondo, A.
    Volume 3, no. 2, e00389-15, 2015. Page 1, column 1, lines 26 and 27: "silencing mediator for retinoic acid and thyroid hormone receptor" should read "singlemolecule real-time."
    American Society for Microbiology, 2017, Genome Announcements, 5(5) (5), English
    [Refereed]
    Scientific journal

  • Nishimura, Y., Ezawa, R., Ishii, J., Ogino, C., Kondo, A.
    The expression of epidermal growth factor receptor (EGFR) across a wide range of tumor cells has attracted attention for use as a tumor marker in drug delivery systems. Therefore, binding molecules with the ability to target EGFR have been developed. Among them, we focused on affibodies that are binding proteins derived from staphylococcal protein A. By displaying affibody (Z(EGFR)) binding to EGFR on the surface of a bio-nanocapsule (BNC) derived from a hepatitis B virus (HBV), we developed an altered BNC (Z(EGFR)-BNC) with a high specificity to EGFR-expressing cells. We considered two different types of Z(EGFR) (Z955 and Z1907), and found that the Z1907 dimer-displaying BNC ([Z1907[(2)-BNC) could effectively bind to EGFR-expressing cells and deliver drugs to the cytosol. Since this study showed that [Z1907](2)-BNC could target EGFR-expressing cells, we would use this particle as a drug delivery carrier for various cancer cells expressing EGFR. (C) 2016 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license.
    PERGAMON-ELSEVIER SCIENCE LTD, Jan. 2017, Bioorganic and Medicinal Chemistry Letters, 27(2) (2), 336 - 341, English
    [Refereed]
    Scientific journal

  • Fukutani, Y., Ishii, J., Kondo, A., Ozawa, T., Matsunami, H., Yohda, M.
    The budding yeast Saccharomyces cerevisiae is equipped with G protein-coupled receptors (GPCR). Because the yeast GPCR signaling mechanism is partly similar to that of the mammalian system, S. cerevisiae can be used for a host of mammalian GPCR expression and ligand-mediated activation assays. However, currently available yeast systems require several hours to observe the responses because they depend on the expression of reporter genes. In this study, we attempted to develop a simple GPCR assay system using split luciferase and -arrestin, which are independent of the endogenous S. cerevisiae GPCR signaling pathways. We applied the split luciferase complementation assay method to S. cerevisiae and found that it can be used to analyze the ligand response of the human somatostatin receptor in S. cerevisiae. On the contrary, the response of the pheromone receptor Ste2 was not observed by the assay. Thus, the split luciferase complementation should be free from the effect of the endogenous GPCR signaling. Biotechnol. Bioeng. 2017;114: 1354-1361. (c) 2017 Wiley Periodicals, Inc.
    WILEY, Jun. 2017, Biotechnology and Bioengineering, 114(6) (6), 1354 - 1361, English
    [Refereed]
    Scientific journal

  • Morita, K., Nomura, Y., Ishii, J., Matsuda, F., Kondo, A., Shimizu, H.
    Bacterial phosphoenol pyruvate carboxylase (PPC) and enzymes in the Entner-Doudoroff (ED) pathway were heter-ologously expressed in Saccharomyces cerevisiae to improve the NADPH supply required for the bio-production of chemicals such as isobutanol. The heterologous expression of PPC from Synechocystis sp. PCC6803 increased in the isobutabol titer 1.45-fold (93.2 +/- 1.6 mg/L) in metabolically engineered S. cerevisiae strains producing isobutanol. This result suggested that the pyruvate and NADPH supply for isobutanol biosynthesis was activated by PPC overexpression. On the other hand, the expression of two enzymes organizing the ED pathway (6-phosphogluconate dehydratase [6PGD] and 2-dehydro-3-deoxy-phosphogluconate aldolase [ICDPGA]) had no effect to isobutabol bio-production. Further analysis, however, revealed that additional expression of 6PGD and KDPGA improved the growth rate of S. cerevisiae strain BY4742 gnd1 Delta. A C-13-labeling experiment using [1 - C-13] glucose also suggested that metabolic flow levels in the ED pathway increased slightly with the additional expression. These results showed that the ED pathway was successfully constructed in S. cerevisiae, even though activity of the pathway was too weak to improve isobutanol biosynthesis. (C) 2017, The Society for Biotechnology, Japan. All rights reserved.
    SOC BIOSCIENCE BIOENGINEERING JAPAN, Sep. 2017, Journal of Bioscience and Bioengineering, 124(3) (3), 263 - 270, English
    [Refereed]
    Scientific journal

  • Hara, K.Y., Kobayashi, J., Yamada, R., Sasaki, D., Kuriya, Y., Hirono-Hara, Y., Ishii, J., Araki, M., Kondo, A.
    Biomass resources are attractive carbon sources for bioproduction because of their sustainability. Many studies have been performed using biomass resources to produce sugars as carbon sources for cell factories. Expression of biomass hydrolyzing enzymes in cell factories is an important approach for constructing biomass-utilizing bioprocesses because external addition of these enzymes is expensive. In particular, yeasts have been extensively engineered to be cell factories that directly utilize biomass because of their manageable responses to many genetic engineering tools, such as gene expression, deletion and editing. Biomass utilizing bioprocesses have also been developed using these genetic engineering tools to construct metabolic pathways. However, sugar input and product output from these cells are critical factors for improving bioproduction along with biomass utilization and metabolic pathways. Transporters are key components for efficient input and output activities. In this review, we focus on transporter engineering in yeast to enhance bioproduction from biomass resources.
    OXFORD UNIV PRESS, Nov. 2017, FEMS Yeast Research, 17(7) (7), fox061, English
    [Refereed]
    Scientific journal

  • Prima, A., Hara, K.Y., Djohan, A.C., Kashiwagi, N., Kahar, P., Ishii, J., Nakayama, H., Okazaki, F., Prasetya, B., Kondo, A., Yopi, Ogino, C.
    This work aims to produce glutathione directly from mannan-based bioresources using engineered Saccharomyces cerevisiae. Mannan proved to be a valuable carbon source for glutathione production by this organism. Mannan-hydrolyzing S. cerevisiae was developed by heterologous expression of mannanase/ mannosidase on its cell surface. This strain efficiently produced glutathione from mannose polysaccharide, beta-1,4-mannan. Furthermore, it produced glutathione from locust bean gum (LBG), a highly dense and inexpensive mannan-based bioresource, as sole carbon source. Glutathione productivity from LBG was enhanced by engineering the glutathione metabolism of mannan-hydrolyzing S. cerevisiae. Expression of extracellular mannanase/mannosidase protein combined with intracellular metabolic engineering is potentially applicable to the efficient, environmentally friendly bioproduction of targeted products from mannan-based bioresources. (C) 2017 Elsevier Ltd. All rights reserved.
    ELSEVIER SCI LTD, Dec. 2017, Bioresource Technology, 245, 1400 - 1406, English
    [Refereed]
    Scientific journal

  • Kaishima, M., Ishii, J., Matsuno, T., Fukuda, N., Kondo, A.
    Green fluorescent protein (GFP), which was originally isolated from jellyfish, is a widely used tool in biological research, and homologs from other organisms are available. However, researchers must determine which GFP is the most suitable for a specific host. Here, we expressed GFPs from several sources in codon-optimized and non-codon-optimized forms in the yeast Saccharomyces cerevisiae, which represents an ideal eukaryotic model. Surprisingly, codon-optimized mWasabi and mNeonGreen, which are typically the brightest GFPs, emitted less green fluorescence than did the other five codon-optimized GFPs tested in S. cerevisiae. Further, commercially available GFPs that have been optimized for mammalian codon usage (e.g., EGFP, AcGFP1 and TagGFP2) unexpectedly exhibited extremely low expression levels in S. cerevisiae. In contrast, codon-optimization of the GFPs for S. cerevisiae markedly increased their expression levels, and the fluorescence intensity of the cells increased by a maximum of 101-fold. Among the tested GFPs, the codon-optimized monomeric mUkG1 from soft coral showed the highest levels of both expression and fluorescence. Finally, the expression of this protein as a fusion-tagged protein successfully improved the reporting system's ability to sense signal transduction and protein-protein interactions in S. cerevisiae and increased the detection rates of target cells using flow cytometry.
    Corresponding, NATURE PUBLISHING GROUP, Oct. 2016, Scientific Reports, 6, 35932, English, International magazine
    [Refereed]
    Scientific journal

  • Nakamura, Y., Hashimoto, T., Ishii, J., Kondo, A.
    G-protein-coupled receptors (GPCRs) are physiologically important membrane proteins that represent major molecular targets in pharmaceutical and medicinal fields. Many GPCRs have been shown to form not only homodimers, but also heterodimers that can confer large functional and physiological diversity and are therefore expected to offer new opportunities for the discovery of new drugs. Yeast is a useful host organism that can be used to investigate the interactions between eukaryotic protein pairs, as demonstrated by the yeast two-hybrid (Y2H) system. Previously, we established reporter gene assay systems to screen for GPCR dimer pairs based on the splitubiquitin membrane Y2H (mY2H) system. However, conventional systems only induce reporter gene expressions from the OFF to ON states. In this study, we therefore designed a reporter switching system that can switch the expressions between two reporter genes (one from ON to OFF and the other from OFF to ON) in response to the Y2H readout. To invoke reporter switching, we took advantage of Cre/loxP site-specific recombination. Through optimization of Cre-mediated reporter gene recombination using the split-ubiquitin mY2H system, we built a dual-color reporter switching system to discern the formations of GPCR dimers. This system enabled monitoring of the formations of homodimers and heterodimers of human serotonin 1A receptor or beta(2)-adrenergic receptor as well as homodimers of the yeast endogenous pheromone receptor (Ste2p) in yeast cells. Our reporter switching system may be a useful tool for identifying potential molecular targets among GPCR dimers, and is also applicable to other reporter gene assay systems. (C) 2016 Wiley Periodicals, Inc.
    Corresponding, WILEY-BLACKWELL, Oct. 2016, Biotechnology and Bioengineering, 113(10) (10), 2178 - 2190, English, International magazine
    [Refereed]
    Scientific journal

  • Inokuma, K., Ishii, J., Hara, K.Y., Mochizuki, M., Hasunuma, T., Kondo, A.
    We determined the genome sequence of the thermotolerant yeast Kluyveromyces marxianus strain NBRC1777. The genome of strain NBRC1777 is composed of 4,912 open reading frames (ORFs) on 8 chromosomes, with a total size of 10,895,581 bp, including mitochondrial DNA.
    American Society for Microbiology, 2016, Genome Announcements, 3(2) (2), e00389 - 15, English
    [Refereed]
    Scientific journal

  • Tsuge, Y., Kudou, M., Kawaguchi, H., Ishii, J., Hasunuma, T., Kondo, A.
    Lignocellulosic hydrolysates contain compounds that inhibit microbial growth and fermentation, thereby decreasing the productivity of biofuel and biochemical production. In particular, the heterocyclic aldehyde furfural is one of the most toxic compounds found in these hydrolysates. We previously demonstrated that Corynebacterium glutamicum converts furfural into the less toxic compounds furfuryl alcohol and 2-furoic acid. To date, however, the genes involved in these oxidation and reduction reactions have not been identified in the C. glutamicum genome. Here, we show that Cgl0331 (designated FudC) is mainly responsible for the reduction of furfural into furfuryl alcohol in C. glutamicum. Deletion of the gene encoding FudC markedly diminished the in vivo reduction of furfural to furfuryl alcohol. Purified His-tagged FudC protein from Escherichia coli was also shown to convert furfural into furfuryl alcohol in an in vitro reaction utilizing NADPH, but not NADH, as a cofactor. Kinetic measurements demonstrated that FudC has a high affinity for furfural but has a narrow substrate range for other aldehydes compared to the protein responsible for furfural reduction in E. coli.
    SPRINGER, Mar. 2016, Applied Microbiology and Biotechnology, 100(6) (6), 2685 - 2692, English
    [Refereed]
    Scientific journal

  • Hattan, J., Shindo, K., Ito, T., Shibuya, Y., Watanabe, A., Tagaki, C., Ohno, F., Sasaki, T., Ishii, J., Kondo, A., Misawa, N.
    A novel terpene synthase ( Tps ) gene isolated from Camellia brevistyla was identified as hedycaryol synthase, which was shown to be expressed specifically in flowers. Camellia plants are very popular because they bloom in winter when other plants seldom flower. Many ornamental cultivars of Camellia have been bred mainly in Japan, although the fragrance of their flowers has not been studied extensively. We analyzed floral scents of several Camellia cultivars by gas chromatography-mass spectrometry (GC-MS) and found that Camellia brevistyla produced various sesquiterpenes in addition to monoterpenes, whereas Camellia japonica and its cross-lines produced only monoterpenes, including linalool as the main product. From a flower of C. brevistyla, we isolated one cDNA encoding a terpene synthase (TPS) comprised of 554 amino acids, which was phylogenetically positioned to a sole gene clade. The cDNA, designated CbTps1, was expressed in mevalonate-pathway-engineered Escherichia coli, which carried the Streptomyces mevalonate-pathway gene cluster in addition to the acetoacetate-CoA ligase gene. A terpene product was purified from recombinant E. coli cultured with lithium acetoacetate, and analyzed by H-1-nulcear magnetic resonance spectroscopy (H-1-NMR) and GC-MS. It was shown that a sesquiterpene hedycaryol was produced, because H-1-NMR signals of the purified product were very broad, and elemol, a thermal rearrangement product from hedycaryol, was identified by GC-MS analysis. Spectroscopic data of elemol were also determined. These results indicated that the CbTps1 gene encodes hedycaryol synthase. Expression analysis of CbTps1 showed that it was expressed specifically in flowers, and hedycaryol is likely to be one of the terpenes that attract insects for pollination of C. brevistyla. A linalool synthase gene, which was isolated from a flower of Camellia saluenensis, is also described.
    SPRINGER, Apr. 2016, Planta, 243(4) (4), 959 - 972, English
    [Refereed]
    Scientific journal

  • Nakamura, Y., Ishii, J., Kondo, A.
    G-protein-coupled receptors (GPCRs) are physiologically important transmembrane proteins that sense signaling molecules such as hormones, neurotransmitters, and various sensory stimuli; GPCRs represent major molecular targets for drug discovery. Although GPCRs traditionally have been thought to function as monomers or homomers, in the recent years these proteins have also been shown to function as heteromers. Heteromerization among GPCRs is expected to generate potentially large functional and physiological diversity and to provide new opportunities for drug discovery. However, due to the existence of numerous combinations, the larger universe of possible GPCR heteromers is unknown, and thus its functional significance is still poorly understood. The oligomerization of GPCRs in living cells now has been demonstrated in mammalian cells and in native tissues by using genetic, biochemical, and physiological approaches, as well as various resonance energy transfer (RET) technologies. In addition, the yeast Saccharomyces cerevisiae, which can serve as a biosensor for monitoring eukaryotic biological processes, can also be used for the identification of functionally significant heteromer pairs of GPCRs. In this review, we focus on studies of GPCR oligomers, and summarize the technologies used to evaluate GPCR oligomerization. We additionally consider the potential limitations of these methods at present, and envision the possible future applications of these techniques.
    BENTHAM SCIENCE PUBL LTD, 2016, Current Medicinal Chemistry, 23(16) (16), 1638 - 1656, English
    [Refereed]
    Scientific journal

  • Shen, L., Nishimura, Y., Matsuda, F., Ishii, J., Kondo, A.
    2-Phenylethanol (2-PE) is a higher aromatic alcohol that is used in the cosmetics and food industries. The budding yeast Saccharomyces cerevisiae is considered to be a suitable host for the industrial production of higher alcohols, including 2-PE. To produce 2-PE from glucose in S. cerevisiae, we searched for suitable 2-keto acid decarboxylase (KDC) and alcohol dehydrogenase (ADH) enzymes of the Ehrlich pathway for overexpression in strain YPH499, and found that overexpression of the ARO10 and/or ADH1 genes increased 2-PE production from glucose. Further, we screened ten BY4741 single-deletion mutants of genes involved in the competing pathways for 2-PE production, and found that strains aro8 Delta and aat2 Delta displayed increased 2-PE production. Based on these results, we engineered a BY4741 strain that overexpressed ARO10 and contained an aro8 Delta deletion, and demonstrated that the strain produced 96 mg/L 2-PE from glucose as the sole carbon source. As this engineered S. cerevisiae strain showed a significant increase in 2-PE production from glucose without the addition of an intermediate carbon substrate, it is a promising candidate for the large-scale production of 2-PE. (C) 2015 The Society for Biotechnology, Japan. All rights reserved.
    SOC BIOSCIENCE BIOENGINEERING JAPAN, Jul. 2016, Journal of Bioscience and Bioengineering, 122(1) (1), 34 - 39, English
    [Refereed]
    Scientific journal

  • Ishii, J., Okazaki, F., Djohan, A. C., Hara, K. Y., Asai-Nakashima, N., Teramura, H., Andriani, A., Tominaga, M., Wakai, S., Kahar, P., Prasetya, Y. B., Ogino, C., Kondo, A.
    Background: Mannans represent the largest hemicellulosic fraction in softwoods and also serve as carbohydrate stores in various plants. However, the utilization of mannans as sustainable resources has been less advanced in sustainable biofuel development. Based on a yeast cell surface-display technology that enables the immobilization of multiple enzymes on the yeast cell walls, we constructed a recombinant Saccharomyces cerevisiae strain that co-displays beta-mannanase and beta-mannosidase; this strain is expected to facilitate ethanol fermentation using mannan as a biomass source. Results: Parental yeast S. cerevisiae assimilated mannose and glucose as monomeric sugars, producing ethanol from mannose. We constructed yeast strains that express tethered beta-mannanase and beta-mannosidase; co-display of the two enzymes on the cell surface was confirmed by immunofluorescence staining and enzyme activity assays. The constructed yeast cells successfully hydrolyzed 1,4-beta-D-mannan and produced ethanol by assimilating the resulting mannose without external addition of enzymes. Furthermore, the constructed strain produced ethanol from 1,4-beta-D-mannan continually during the third batch of repeated fermentation. Additionally, the constructed strain produced ethanol from ivory nut mannan; ethanol yield was improved by NaOH pretreatment of the substrate. Conclusions: We successfully displayed beta-mannanase and beta-mannosidase on the yeast cell surface. Our results clearly demonstrate the utility of the strain co-displaying beta-mannanase and beta-mannosidase for ethanol fermentation from mannan biomass. Thus, co-tethering beta-mannanase and beta-mannosidase on the yeast cell surface provides a powerful platform technology for yeast fermentation toward the production of bioethanol and other biochemicals from lignocellulosic materials containing mannan components.
    BIOMED CENTRAL LTD, Sep. 2016, Biotechnology for Biofuels, 9(1) (1), 188, English
    [Refereed]
    Scientific journal

  • Yamamoto, K., Hara, K. Y., Morita, T., Nishimura, A., Sasaki, D., Ishii, J., Ogino, C., Kizaki, N., Kondo, A.
    Background: Red yeast, Xanthophyllomyces dendrorhous is the only yeast known to produce astaxanthin, an antioxidant isoprenoid (carotenoid) widely used in the aquaculture, food, pharmaceutical and cosmetic industries. The potential of this microorganism as a platform cell factory for isoprenoid production has been recognized because of high flux through its native terpene pathway. Recently, we developed a multiple gene expression system in X. dendrorhous and enhanced the mevalonate synthetic pathway to increase astaxanthin production. In contrast, the mevalonate synthetic pathway is suppressed by ergosterol through feedback inhibition. Therefore, releasing the mevalonate synthetic pathway from this inhibition through the deletion of genes involved in ergosterol synthesis is a promising strategy to improve isoprenoid production. An efficient method for deleting diploid genes in X. dendrorhous, however, has not yet been developed. Results: Xanthophyllomyces dendrorhous was cultivated under gradually increasing concentrations of antibiotics following the introduction of antibiotic resistant genes to be replaced with target genes. Using this method, double CYP61 genes encoding C-22 sterol desaturases relating to ergosterol biosynthesis were deleted sequentially. This double CYP61 deleted strain showed decreased ergosterol biosynthesis compared with the parental strain and single CYP61 disrupted strain. Additionally, this double deletion of CYP61 genes showed increased astaxanthin production compared with the parental strain and the single CYP61 knockout strain. Finally, astaxanthin production was enhanced by 1.4-fold compared with the parental strain, although astaxanthin production was not affected in the single CYP61 knockout strain. Conclusions: In this study, we developed a system to completely delete target diploid genes in X. dendrorhous. Using this method, we deleted diploid CYP61 genes involved in the synthesis of ergosterol that inhibits the pathway for mevalonate, which is a common substrate for isoprenoid biosynthesis. The resulting decrease in ergosterol biosynthesis increased astaxanthin production. The efficient method for deleting diploid genes developed in this study has the potential to improve industrial production of various isoprenoids in X. dendrorhous.
    BIOMED CENTRAL LTD, Sep. 2016, Microbial Cell Factories, 15(1) (1), 155, English
    [Refereed]
    Scientific journal

  • Inokuma, K., Bamba, T., Ishii, J., Ito, Y., Hasunuma, T., Kondo, A.
    Recombinant yeast strains displaying aheterologous cellulolytic enzymes on their cell surfaces using a glycosylphosphatidylinositol (GPI) anchoring system are a promising strategy for bioethanol production from lignocellulosic materials. A crucial step for cell wall localization of the enzymes is the intracellular transport of proteins in yeast cells. Therefore, the addition of a highly efficient secretion signal sequence is important to increase the amount of the enzymes on the yeast cell surface. In this study, we demonstrated the effectiveness of a novel signal peptide (SP) sequence derived from the Saccharomyces cerevisiae SED1 gene for cell-surface display and secretory production of cellulolytic enzymes. Gene cassettes with SP sequences derived fromS. cerevisiae SED1 (SED1SP), Rhizopus oryzae glucoamylase (GLUASP), and S. cerevisiae alpha-mating pheromone (MF alpha 1SP) were constructed for cell-surface display of Aspergillus aculeatus beta-glucosidase (BGL1) and Trichoderma reesei endoglucanase II (EGII). These gene cassettes were integrated into the S. cerevisiae genome. The recombinant strains with the SED1SP showed higher cell-surface BGL and EG activities than those with the conventional SP sequences (GLUASP and MF alpha 1SP). The novel SP sequence also improved the secretory production of BGL and EG in S. cerevisiae. The extracellular BGL activity of the recombinant strains with the SED1SP was 1.3- and 1.9-fold higher than the GLUASP and MF alpha 1SP strains, respectively. Moreover, the utilization of SED1SP successfully enhanced the secretory production of BGL in Pichia pastoris. The utilization of the novel SP sequence is a promising option for highly efficient cell-surface display and secretory production of heterologous proteins in various yeast species. (C) 2016 Wiley Periodicals, Inc.
    WILEY-BLACKWELL, Nov. 2016, Biotechnology and Bioengineering, 113(11) (11), 2358 - 2366, English
    [Refereed]
    Scientific journal

  • Fukuda, N., Kaishima, M., Ishii, J., Kondo, A., Honda, S.
    Sake yeasts belong to the budding yeast species Saccharomyces cerevisiae and have high fermentation activity and ethanol production. Although the traditional crossbreeding of sake yeasts is a time-consuming and inefficient process due to the low sporulation rates and spore viability of these strains, considerable effort has been devoted to the development of hybrid strains with superior brewing characteristics. In the present work, we describe a growth selection system for a- and α-type cells aimed at the crossbreeding of industrial yeasts, and performed hybridizations with sake yeast strains Kyokai No. 6, No. 7 and No. 9 to examine the feasibility of this approach. We successfully generated both a- and α-type strains from all parental strains, and acquired six types of hybrids by outcrossing. One of these hybrid strains was subjected to continuous crossbreeding, yielding the multi-hybrid strain, which inherited the genetic characteristics of Kyokai No. 6, No. 7 and No. 9. Notably, because all of the genetic modifications of the yeast cells were introduced using plasmids, these traits can be easily removed. The approach described here has the potential to markedly accelerate the crossbreeding of industrial yeast strains with desirable properties.
    Springer Verlag, Dec. 2016, Amb Express, 6(1) (1), 45, English
    [Refereed]
    Scientific journal

  • 竹村 秀史, 鈴木 宗典, 原田 尚志, 三沢 典彦, 石井 純, 近藤 昭彦, 播本 孝史
    (公社)日本生物工学会, Sep. 2015, 日本生物工学会大会講演要旨集, 平成27年度, 337 - 337, Japanese

  • Suzuki H, Ishii J, Kondo A, Yoshida Ki
    Microbes employ cell membranes for reducing exogenous stresses. Polyamino acid display on microbial cell surfaces and their effects on microbial chemical stress tolerance were examined. Growth analysis revealed that displays of polyarginine, polyaspartate and polytryptophan substantially enhanced tolerance of Escherichia coli to NaCl. A titration assay indicated that polyarginine and polyaspartate altered cell surface charges, implying tolerance enhancement via ion atmosphere and/or ionic bond network formations for electrostatic ion repulsion. The enhancement by polytryptophan may have arisen from surface hydrophobicity increase for hydrophobic ion exclusion, because of a strong correlation between hydrophobic characters of amino acids and their effects on tolerance enhancement. The display also enhanced tolerance to other salts and/or alcohols in E. coli and to NaCl in Saccharomyces cerevisiae. Thus polyamino acid display has the potential as an approach for conferring chemical stress tolerance on various microbes.
    SPRINGER, Feb. 2015, Biotechnology letters., 37(2) (2), 429 - 435, English
    [Refereed]
    Scientific journal


  • HASUNUMA Tomohisa, ISHII Jun, OGINO Chiaki, KONDO Akihiko
    持続可能な社会へ向かうためには再生可能エネルギーが中心的な役割を果たすことが求められている.そのなかで,バイオマスから液体燃料やバルクケミカルを経済性良く,高効率で生産する技術の開発が期待されている.バイオマスとしては,安定的な供給が可能で,食糧と競合しないリグノセルロース系バイオマスの利活用が望まれている.本稿ではリグノセルロース系バイオマスからのエタノールの製造プロセスについて研究の課題と最新の知見を紹介するとともに,バイオプロセスによるバルクケミカル生産に関する最近の研究例についても紹介する.
    Japan Society for Bioscience, Biotechnology, and Agrochemistry, 2015, Kagaku To Seibutsu, 53(10) (10), 689 - 695, Japanese

  • Hasunuma, T., Ishii, J., Kondo, A.
    Conferring biomass hydrolysis activity on yeast through genetic engineering has paved the way for the development of groundbreaking processes for producing liquid fuels and commodity chemicals from lignocellulosic biomass. However, the overproduction and misfolding of heterologous and endogenous proteins can trigger cellular stress, increasing the metabolic burden and retarding growth. Improving the efficiency of lignocellulosic breakdown requires engineering of yeast secretory pathway based on system-wide metabolic analysis as well as DNA constructs for enhanced cellulase gene expression with advanced molecular biology tools. Also, yeast is subjected to severe stress due to toxic compounds generated during lignocellulose pretreatment in consolidated saccharification and fermentation processes. The prospect for development of robust yeast strains makes combining evolutionary and rational engineering strategies.
    ELSEVIER SCI LTD, Dec. 2015, Current Opinion in Chemical Biology, 29, 1 - 9, English
    [Refereed]
    Scientific journal

  • Kaishima, M., Ishii, J., Fukuda, N., Kondo, A.
    Protein-protein interactions (PPIs) are crucial for the vast majority of biological processes. We previously constructed a G gamma recruitment system to screen PPI candidate proteins and desirable affinity-altered (affinity-enhanced and affinity-attenuated) protein variants. The methods utilized a target protein fused to a mutated G-protein gamma subunit (G gamma(cyto)) lacking the ability to localize to the inner leaflet of the plasma membrane. However, the previous systems were adapted to use only soluble cytosolic proteins as targets. Recently, membrane proteins have been found to form the principal nodes of signaling involved in diseases and have attracted a great deal of interest as primary drug targets. Here, we describe new protocols for the G gamma recruitment systems that are specifically designed to use membrane proteins as targets to overcome previous limitations. These systems represent an attractive approach to exploring novel interacting candidates and affinity-altered protein variants and their interactions with proteins on the inner side of the plasma membrane, with high specificity and selectivity.
    NATURE PUBLISHING GROUP, Nov. 2015, Scientific Reports, 5, 16723, English
    [Refereed]
    Scientific journal

  • Ida, K., Ishii, J., Matsuda, F., Kondo, T., Kondo, A.
    Background: Isobutanol is an important biorefinery target alcohol that can be used as a fuel, fuel additive, or commodity chemical. Baker's yeast, Saccharomyces cerevisiae, is a promising organism for the industrial manufacture of isobutanol because of its tolerance for low pH and resistance to autolysis. It has been reported that gene deletion of the pyruvate dehydrogenase complex, which is directly involved in pyruvate metabolism, improved isobutanol production by S. cerevisiae. However, the engineering strategies available for S. cerevisiae are immature compared to those available for bacterial hosts such as Escherichia coli, and several pathways in addition to pyruvate metabolism compete with isobutanol production. Results: The isobutyrate, pantothenate or isoleucine biosynthetic pathways were deleted to reduce the outflow of carbon competing with isobutanol biosynthesis in S. cerevisiae. The judicious elimination of these competing pathways increased isobutanol production. ILV1 encodes threonine ammonia-lyase, the enzyme that converts threonine to 2-ketobutanoate, a precursor for isoleucine biosynthesis. S. cerevisiae mutants in which ILV1 had been deleted displayed 3.5-fold increased isobutanol productivity. The Delta ILV1 strategy was further combined with two previously established engineering strategies (activation of two steps of the Ehrlich pathway and the transhydrogenase-like shunt), providing 11-fold higher isobutanol productivity as compared to the parent strain. The titer and yield of this engineered strain was 224 +/- 5 mg/L and 12.04 +/- 0.23 mg/g glucose, respectively. Conclusions: The deletion of competitive pathways to reduce the outflow of carbon, including ILV1 deletion, is an important strategy for increasing isobutanol production by S. cerevisiae.
    BIOMED CENTRAL LTD, Apr. 2015, Microbial Cell Factories, 14, 62, English
    [Refereed]
    Scientific journal

  • Suffian, I. F. B., Nishimura, Y., Morita, K., Nakamura-Tsuruta, S., Al-Jamal, K. T., Ishii, J., Ogino, C., Kondo, A.
    Background: The hepatitis B virus core (HBc) particle is known as a promising new carrier for the delivery of drugs and nucleic acids. However, since the arginine-rich domain that is located in the C-terminal region of the HBc monomer binds to the heparan sulphate proteoglycan on the cell surface due to its positive charge, HBc particles are introduced non-specifically into a wide range of cells. To avoid non-specific cellular uptake with the intent to control the ability of cell targeting, we individually replaced the respective arginine (R) residues of the arginine-rich domain located in amino acid positions 150-159 in glycine (G) residues. Results: The mutated HBc particles in which R154 was replaced with glycine (G) residue (R154G) showed a drastic decrease in the ability to bind to the heparan sulphate proteoglycan and to avoid non-specific cellular uptake by several types of cancer cells. Conclusions: Because this mutant particle retains most of its C-terminal arginine-rich residues, it would be useful in the targeting of specificity-altered HBc particles in the delivery of nucleic acids.
    BIOMED CENTRAL LTD, Feb. 2015, Journal of Nanobiotechnology, 13(1) (1), 15, English
    [Refereed]
    Scientific journal

  • Fukutani, Y., Hori, A., Tsukada, S., Sato, R., Ishii, J., Kondo, A., Matsunami, H., Yohda, M.
    Olfaction depends on the selectivity and sensitivity of olfactory receptors. Previous attempts at constructing a mammalian olfactory receptor-based artificial odorant sensing system in the budding yeast Saccharomyces cerevisiae suffered from low sensitivity and activity. This result may be at least in part due to poor functional expression of olfactory receptors and/or limited solubility of some odorants in the medium. In this study, we examined the effects of two types of accessory proteins, receptor transporting protein 1 short and odorant binding proteins, in improving odor-mediated activation of olfactory receptors expressed in yeast. We found that receptor transporting protein 1 short enhanced the membrane expression and ligand-induced responses of some olfactory receptors. Coexpression of odorant binding proteins of the silkworm moth Bombyx mori enhanced the sensitivity of a mouse olfactory receptor. Our results suggest that different classes of accessory proteins can confer sensitive and robust responses of olfactory receptors expressed in yeast. Inclusion of accessory proteins may be essential in the future development of practical olfactory receptor-based odorant sensors. (C) 2014 Elsevier Inc. All rights reserved.
    ACADEMIC PRESS INC ELSEVIER SCIENCE, Feb. 2015, Analytical Biochemistry, 471, 1 - 8, English
    [Refereed]
    Scientific journal

  • Inokuma, K., Yoshida, T., Ishii, J., Hasunuma, T., Kondo, A.
    Recombinant yeast strains that display heterologous amylolytic enzymes on their cell surface via the glycosylphosphatidylinositol (GPI)-anchoring system are considered as promising biocatalysts for direct ethanol production from starchy materials. For the effective hydrolysis of these materials, the ratio optimization of multienzyme activity displayed on the cell surface is important. In this study, we have presented a ratio control system of multienzymes displayed on the yeast cell surface by using different GPI-anchoring domains. The novel gene cassettes for the cell-surface display of Streptococcus bovis alpha-amylase and Rhizopus oryzae glucoamylase were constructed using the Saccharomyces cerevisiae SED1 promoter and two different GPI-anchoring regions derived from Saccharomyces cerevisiae SED1 or SAG1. These gene cassettes were integrated into the Saccharomyces cerevisiae genome in different combinations. Then, the cell-surface alpha-amylase and glucoamylase activities and ethanol productivity of these recombinant strains were evaluated. The combinations of the gene cassettes of these enzymes affected the ratio of cell-surface alpha-amylase and glucoamylase activities and ethanol productivity of the recombinant strains. The highest ethanol productivity from raw starch was achieved by the strain harboring one alpha-amylase gene cassette carrying the SED1-anchoring region and two glucoamylase gene cassettes carrying the SED1-anchoring region (BY-AASS/GASS/GASS). This strain yielded 22.5 +/- 0.6 g/L of ethanol from 100 g/L of raw starch in 120 h of fermentation.
    SPRINGER, Feb. 2015, Applied Microbiology and Biotechnology, 99(4) (4), 1655 - 1663, English
    [Refereed]
    Scientific journal

  • Takenaka, M., Miyachi, Y., Ishii, J., Ogino, C., Kondo, A.
    An atomic force microscope (AFM) can measure the adhesion force between a sample and a cantilever while simultaneously applying a rupture force during the imaging of a sample. An AFM should be useful in targeting specific proteins on a cell surface. The present study proposes the use of an AFM to measure the adhesion force between targeting receptors and their ligands, and to map the targeting receptors. In this study, Ste2p, one of the G protein-coupled receptors (GPCRs), was chosen as the target receptor. The specific force between Ste2p on a yeast cell surface and a cantilever modified with its ligand, a-factor, was measured and found to be approximately 250 pN. In addition, through continuous measuring of the cell surface, a mapping of the receptors on the cell surface could be performed, which indicated the differences in the Ste2p expression levels. Therefore, the proposed AFM system is accurate for cell diagnosis.
    ROYAL SOC CHEMISTRY, 2015, Nanoscale, 7(11) (11), 4956 - 4963, English
    [Refereed]
    Scientific journal

  • Nakamura, Y., Ishii, J., Kondo, A.
    The monoamine neurotransmitter serotonin (5-HT) regulates a wide spectrum of human physiology through the 5-HT receptor family. One such receptor, the 5-HT1A receptor (HTR1A), is the most widely studied subtype and represents a significant molecular target in medicinal and therapeutic fields. Yeast-based fluorescent reporter systems have proven to be especially useful for GPCR assays, since detection using a fluorescent reporter considerably simplifies measurement procedures. However, previously reported systems using enhanced green fluorescent protein (EGFP) as the reporter in yeast still showed low signal-to-noise (S/N) ratios, making EGFP difficult to apply as an easily accessible tool. Therefore, we constructed a refined yeast-based GPCR biosensor employing a high-sensitivity strain that incorporated both a G-engineered receptor and a fluorescent reporter (ZsGreen). As we report here, the refined yeast-based fluorescent biosensor was applied successfully to antagonist characterization and analysis of site-directed mutants of the HTR1A receptor. Pindolol, a known antagonist of HTR1A, specifically inhibited agonist-induced signaling, demonstrating the ease of evaluating inhibition effects using our reporter strain. Characterization of site-specific receptor mutants confirmed the role of specific targeted residues, including the highly conserved DRY motif, in the activation of HTR1A. Thus, our refined yeast biosensor strain, which incorporates a ZsGreen reporter and an engineered G receptor, is expected to serve as a simple and practical sensing tool for evaluating the ligand candidates and defining residues important to the function of human GPCRs. Biotechnol. Bioeng. 2015;112: 1906-1915. (c) 2015 Wiley Periodicals, Inc.
    WILEY-BLACKWELL, Sep. 2015, Biotechnology and Bioengineering, 112(9) (9), 1906 - 1915, English
    [Refereed]
    Scientific journal

  • Hiraoka, R., Funasaki, Y., Ishii, J., Maruyama, T.
    A novel degradable polyanion, poly(phthalic ethylene glycol ester), was synthesized in one pot in a single step. The degradable polyanion assembles with various polycations to form layer-by-layer films that can encapsulate physiologically active biomolecules. Polyanion degradation can induce film disassembly and release of the encapsulated functional protein.
    ROYAL SOC CHEMISTRY, 2015, Chemical Communications, 51(98) (98), 17447 - 17450, English
    [Refereed]
    Scientific journal

  • 竹村 秀史, 鈴木 宗典, 梅野 太輔, 原田 尚志, 三沢 典彦, 石井 純, 近藤 昭彦, 播本 孝史
    (公社)日本生物工学会, Aug. 2014, 日本生物工学会大会講演要旨集, 平成26年度, 161 - 161, Japanese

  • Signaling assays for detection of human G-protein-coupled receptors in yeast
    Yasuyuki Nakamura, Jun Ishii, Akihiko Kondo
    Aug. 2014, Bio-protocol, 4(16) (16), e1206, English
    [Refereed]
    Scientific journal

  • Ishii, J., Oda, A., Togawa, S., Fukao, A., Fujiwara, T., Ogino, C., Kondo, A.
    Neurotensin receptor type-I (NTSR1) is a member of the G-protein-coupled receptor (GPCR) family. The natural ligand of NTSR1 is neurotensin (NT), a neuromodulator of the central nervous system. Because NT is also involved in many oncogenic actions, the signaling mediator NTSR1 is a significant molecular target in medicinal and therapeutic fields. In the current study, we constructed a fluorescence-based microbial yeast biosensor that can monitor the activation of human NTSR1 signaling responding to its agonist. To increase the sensitivity of the biosensor, a yeast strain with the green fluorescent protein (GFP) reporter gene was genetically engineered to enhance binding with human NTSRI expressed on the membrane. Following previous reports, the 5 carboxy-terminal amino acid residues of the guanine nucleotide binding protein a-subunit (G alpha) in yeast Gpalp were substituted with the equivalent human G alpha, sequences (Gpal/G alpha(q) transplant). After optimizing the assay conditions, the G alpha-engineered yeast demonstrated significantly improved sensing for NTSR1 signaling. Because detection using a GFP fluorescence reporter considerably simplifies the measurement procedure, this microbial fluorescence sensor holds promise for use in the diagnosis of NTSR1-associated diseases and the development of agonists. (C) 2013 Elsevier Inc. All rights reserved.
    ACADEMIC PRESS INC ELSEVIER SCIENCE, Feb. 2014, Analytical Biochemistry, 446(1) (1), 37 - 43, English
    [Refereed]
    Scientific journal

  • Nakamura, Y., Takemoto, N., Ishii, J., Kondo, A.
    Understanding the role of G-protein-coupled receptor (GPCR) dimerization in cellular function has now become a major research focus. The potentially large functional and physiological diversity of dimerization among GPCRs is expected to provide opportunities for novel drug discovery. However, there is currently a lack of cell-based assays capable of specific profiling for the functional consequences of dimerization linked to ligand-mediated signaling. Here, we present an advanced method to simultaneously analyze the dimerization and ligand response of GPCRs using two yeast-based systems for split-ubiquitin two-hybrid assay and G-protein signaling assay. To permit simultaneous detection, we established a two-color (dual-color) fluorescence reporter gene assay using enhanced green fluorescent protein (EGFP) and a far-red derivative of the tetrameric fluorescent protein DsRed-Express2 (E2-Crimson). In the present study, we tested our method first by analyzing dimerization and ligand-mediated signaling by the yeast endogenous pheromone receptor (Ste2p). Second, we showed that the system facilitated mutational analysis of domains involved in dimerization and signaling by Ste2p. Third, we successfully demonstrated that the system could simultaneously monitor homo- and hetero-dimerization and somatostatin-induced signaling in the test case of the human SSTR5 somatostatin receptor. Our strategy is expected to provide a useful tool for the elucidation of molecular biological functions of GPCR dimers and for the screening of GPCR dimer-specific agonistic ligands. Biotechnol. Bioeng. 2014;111: 586-596. (c) 2013 Wiley Periodicals, Inc.
    WILEY, Mar. 2014, Biotechnology and Bioengineering, 111(3) (3), 586 - 596, English
    [Refereed]
    Scientific journal

  • Nishimura, Y., Takeda, K., Ezawa, R., Ishii, J., Ogino, C., Kondo, A.
    Background: An affibody-displaying bio-nanocapsule (Z(HER2)-BNC) with a hepatocyte specificity derived from hepatitis B virus (HBV) was converted into an affibody, Z(HER2), that recognizes HER2 receptors. This affibody was previously reported to be the result of the endocytosis-dependent specific uptake of proteins and siRNA into target cancer cells. To assist the endosomal escape of inclusions, a helper lipid with pH-sensitive fusogenic ability (1,2-dioleoyl-sn-glycero-3-phos phoethanolamine; DOPE) was conjugated with a Z(HER2)-BNC. Findings: In this study, we displayed a pH-sensitive fusogenic GALA peptide on the surface of a particle in order to confer the ability of endosomal escape to a Z(HER2)-BNC. A GALA-displaying Z(HER2)-BNC purified from yeast uneventfully formed a particle structure. Furthermore, endosomal escape of the particle was facilitated after endocytic uptake and release of the inclusions to the cytoplasm without the cell toxicity. Conclusion: The genetic fusion of a GALA peptide to the virus-like particle confers the ability of endosomal escape.
    BIOMED CENTRAL LTD, Apr. 2014, Journal of Nanobiotechnology, 12(1) (1), 11, English
    [Refereed]
    Scientific journal

  • Ishii, J., Kondo, T., Makino, H., Ogura, A., Matsuda, F., Kondo, A.
    Yeast has the potential to be used in bulk-scale fermentative production of fuels and chemicals due to its tolerance for low pH and robustness for autolysis. However, expression of multiple external genes in one host yeast strain is considerably labor-intensive due to the lack of polycistronic transcription. To promote the metabolic engineering of yeast, we generated systematic and convenient genetic engineering tools to express multiple genes in Saccharomyces cerevisiae. We constructed a series of multi-copy and integration vector sets for concurrently expressing two or three genes in S.cerevisiae by embedding three classical promoters. The comparative expression capabilities of the constructed vectors were monitored with green fluorescent protein, and the concurrent expression of genes was monitored with three different fluorescent proteins. Our multiple gene expression tool will be helpful to the advanced construction of genetically engineered yeast strains in a variety of research fields other than metabolic engineering.
    WILEY-BLACKWELL, May 2014, FEMS Yeast Research, 14(3) (3), 399 - 411, English
    [Refereed]
    Scientific journal

  • Nishimura, Y., Ishii, J., Ogino, C., Kondo, A.
    Techniques using nanotechnology in the detection and treatment of cancers have made great progress in multidisciplinary fields. The advances in drug delivery systems (DDSs) have been supported mainly by the development of varied nanoparticles (NPs). Although the NPs based on organic and inorganic materials are integral parts in DDSs, bio-nanoparticles containing biopolymer and virus-like particles (VLPs) are attractive biomaterials for DDSs because of their unique features originating in bio-based materials, such as biocompatibility, biodegradability and low immunogenicity. It is notable that these NPs additionally have a great advantage to enable the easy and flexible alteration of their features by genetic engineering approaches. Controlling the sequence and oligomeric process of polypeptide genes permits a variety of choices in type or size of biopolymeric NPs (e.g., elastin-like polypeptide NPs). In contrast, the functional genes are often inserted into the coding sequences for self-assembled proteins to give the VLPs (e.g., hemagglutinating virus of Japan, adeno-associated virus, human immunodeficiency virus-1, simian virus 40 and hepatitis B virus) additional functions. Thus, genetic engineering readily allow alterations of the properties of NPs (e.g., particle shape, size and stability) and grant of new abilities (e.g., cell-specificity and drug loading and release). In this review, we introduce recent advances in bio-nanoparticles from the standpoint of engineering.
    AMER SCIENTIFIC PUBLISHERS, Sep. 2014, Journal of Biomedical Nanotechnology, 10(9) (9), 2063 - 2085, English
    [Refereed]
    Scientific journal

  • Kaishima, M., Fukuda, N., Ishii, J., Kondo, A.
    Molecules that can control protein-protein interactions (PPIs) have recently drawn attention as new drug pipeline compounds. Here, we report a technique to screen desirable affinity-altered (affinity-enhanced and affinity-attenuated) protein variants. We previously constructed a screening system based on a target protein fused to a mutated G-protein gamma subunit (G gamma(cyto)) lacking membrane localization ability. This ability, required for signal transmission, is restored by recruiting G gamma(cyto) into the membrane only when the target protein interacts with an artificially membrane-anchored candidate protein, thereby allowing interacting partners (G gamma recruitment system) to be searched and identified. In the present study, the G gamma recruitment system was altered by integrating the cytosolic expression of a third protein as a competitor to set a desirable affinity threshold. This enabled the reliable selection of both affinity-enhanced and affinity-attenuated protein variants. The presented approach may facilitate the development of therapeutic proteins that allow the control of PPIs.
    PUBLIC LIBRARY SCIENCE, Sep. 2014, Plos One, 9(9) (9), e108229, English
    [Refereed]
    Scientific journal

  • Tsuge, Y., Hori, Y., Kudou, M., Ishii, J., Hasunuma, T., Kondo, A.
    The toxic fermentation inhibitors in lignocellulosic hydrolysates raise serious problems for the microbial production of fuels and chemicals. Furfural is considered to be one of the most toxic compounds among these inhibitors. Here, we describe the detoxification of furfural in Corynebacterium glutamicum ATCC13032 under both aerobic and anaerobic conditions. Under aerobic culture conditions, furfuryl alcohol and 2-furoic acid were produced as detoxification products of furfural. The ratio of the products varied depending on the initial furfural concentration. Neither furfuryl alcohol nor 2-furoic acid showed any toxic effect on cell growth, and both compounds were determined to be the end products of furfural degradation. Interestingly, unlike under aerobic conditions, most of the furfural was converted to furfuryl alcohol under anaerobic conditions, without affecting the glucose consumption rate. Both the NADH/NAD(+) and NADPH/NADP(+) ratio decreased in the accordance with furfural concentration under both aerobic and anaerobic conditions. These results indicate the presence of a single or multiple endogenous enzymes with broad and high affinity for furfural and co-factors in C. glutamicum ATCC13032.
    SPRINGER, Oct. 2014, Applied Microbiology and Biotechnology, 98(20) (20), 8675 - 8683, English
    [Refereed]
    Scientific journal

  • Nakamura, Y., Ishii, J., Kondo, A.
    Angiotensin II (Ang II) type 1 receptor (AGTR1) is a G-protein-coupled receptor (GPCR). Its natural ligand, Ang II, is an important effector molecule controlling blood pressure and volume in the cardiovascular system, and is consequently involved in various diseases such as hypertension and heart failure. Thus, the signaling mediator, AGTR1, is a significant molecular target in medicinal and therapeutic fields. Yeast is a useful organism for sensing GPCR signaling because it provides a simplified version of the complicated machinery used by mammalian cells for signal transduction. Although yeast cells can successfully transmit a signal through a variety of human GPCRs expressed in the cell membrane, there have been no reports of the functional activation of AGTR1-mediated signaling in yeast cells. In the present study, we introduced a single mutation into human AGTR1 and used yeast-human chimeric G alpha to exert the functional activation of AGTR1 in yeast cells. The engineered yeast cells expressing AGTR1 mutated at Asn295 and the chimeric G alpha successfully transmitted the signal inside the yeast cells in response to Ang II peptide and its analogs (Ang III and Ang IV peptides) added to the assay medium. Further, we demonstrated that the autocrine Ang II peptide and its analog, produced and secreted by the engineered yeast cells, could by themselves promote AGTR1-mediated signaling. This means that screening for agonistic peptides with various sequences from a self-produced genetic library would be a viable strategy. Thus, the constructed yeast biosensor, integrating an Asn295-mutated AGTR1 receptor, will be valuable in the design of drugs to treat AGTR1-related diseases. (C) 2014 Wiley Periodicals, Inc.
    WILEY-BLACKWELL, Nov. 2014, Biotechnology and Bioengineering, 111(11) (11), 2220 - 2228, English
    [Refereed]
    Scientific journal

  • Kondo, A., Ishii, J., Hara, K.Y., Hasunuma, T., Matsuda, F.
    Synthetic bioengineering is a strategy for developing useful microbial strains with innovative biological functions. Novel functions are designed and synthesized in host microbes with the aid of advanced technologies for computer simulations of cellular processes and the system-wide manipulation of host genomes. Here, we review the current status and future prospects of synthetic bioengineering in the yeast Saccharomyces cerevisiae for bio-refinery processes to produce various commodity chemicals from lignocellulosic biomass. Previous studies to improve assimilation of xylose and production of glutathione and butanol suggest a fixed pattern of problems that need to be solved, and as a crucial step, we now need to identify promising targets for further engineering of yeast metabolism. Metabolic simulation, transcriptomics, and metabolomics are useful emerging technologies for achieving this goal, making it possible to optimize metabolic pathways. Furthermore, novel genes responsible for target production can be found by analyzing large-scale data. Fine-tuning of enzyme activities is essential in the latter stage of strain development, but it requires detailed modeling of yeast metabolic functions. Recombinant technologies and genetic engineering are crucial for implementing metabolic designs into microbes. In addition to conventional gene manipulation techniques, advanced methods, such as multicistronic expression systems, marker-recycle gene deletion, protein engineering, cell surface display, genome editing, and synthesis of very long DNA fragments, will facilitate advances in synthetic bioengineering. (C) 2012 Elsevier B.V. All rights reserved.
    ELSEVIER SCIENCE BV, Jan. 2013, Journal of Biotechnology, 163(2) (2), 204 - 216, English
    [Refereed]
    Scientific journal

  • Suga, H., Matsuda, F., Hasunuma, T., Ishii, J., Kondo, A.
    Three enzymes responsible for the transhydrogenase-like shunt, including malic enzyme (encoded by MAE1), malate dehydrogenase (MDH2), and pyruvate carboxylase (PYC2), were overexpressed to regulate the redox state in xylose-fermenting recombinant Saccharomyces cerevisiae. The YPH499XU/MAE1 strain was constructed by overexpressing native Mae1p in the YPH499XU strain expressing xylose reductase and xylitol dehydrogenase from Scheffersomyces stipitis, and native xylulokinase. Analysis of the xylose fermentation profile under semi-anaerobic conditions revealed that the ethanol yield in the YPH499XU/MAE1 strain (0.38 +/- 0.01 g g(-1) xylose consumed) was improved from that of the control strain (0.31 +/- 0.01 g g(-1) xylose consumed). Reduced xylitol production was also observed in YPH499XU/MAE1, suggesting that the redox balance was altered by Mae1p overexpression. Analysis of intracellular metabolites showed that the redox imbalance during xylose fermentation was partly relieved in the transformant. The specific ethanol production rate in the YPH499XU/MAE1-MDH2 strain was 1.25-fold higher than that of YPH499XU/MAE1 due to the additional overexpression of Mdh2p, whereas the ethanol yield was identical to that of YPH499XU/MAE1. The specific xylose consumption rate was drastically increased in the YPH499XU/MAE1-MDH2-PYC2 strain. However, poor ethanol yield as well as increased production of xylitol was observed. These results demonstrate that the transhydrogenase function implemented in S. cerevisiae can regulate the redox state of yeast cells.
    SPRINGER, Feb. 2013, Applied Microbiology and Biotechnology, 97(4) (4), 1669 - 1678, English
    [Refereed]
    Scientific journal

  • Ishii, J., Yoshimura, K., Hasunuma, T., Kondo, A.
    Several alcohol dehydrogenase (ADH)-related genes have been identified as enzymes for reducing levels of toxic compounds, such as, furfural and/or 5-hydroxymethylfurfural (5-HMF), in hydrolysates of pretreated lignocelluloses. To date, overexpression of these ADH genes in yeast cells have aided ethanol production from glucose or glucose/xylose mixture in the presence of furfural or 5-HMF. However, the effects of these ADH isozymes on ethanol production from xylose as a sole carbon source remain uncertain. We showed that overexpression of mutant NADH-dependent ADH1 derived from TMB3000 strain in the recombinant Saccharomyces cerevisiae, into which xylose reductase (XR) and xylitol dehydrogenase (XDH) pathway of Pichia stipitis has been introduced, improved ethanol production from xylose as a sole carbon source in the presence of 5-HMF. Enhanced furan-reducing activity is able to regenerate NAD+ to relieve redox imbalance, resulting in increased ethanol yield arising from decreased xylitol accumulation. In addition, we found that overexpression of wild-type ADH1 prevented the more severe inhibitory effects of furfural in xylose fermentation as well as overexpression of TMB3000-derived mutant. After 120 h of fermentation, the recombinant strains overexpressing wild-type and mutant ADH1 completely consumed 50 g/L xylose in the presence of 40 mM furfural and most efficiently produced ethanol (15.70 g/L and 15.24 g/L) when compared with any other test conditions. This is the first report describing the improvement of ethanol production from xylose as the sole carbon source in the presence of furan derivatives with xylose-utilizing recombinant yeast strains via the overexpression of ADH-related genes. © 2012 Springer-Verlag.
    Mar. 2013, Applied Microbiology and Biotechnology, 97(6) (6), 2597 - 2607, English
    [Refereed]
    Scientific journal

  • Nishimura, Y., Mimura, W., Mohamed Suffian, I.F., Amino, T., Ishii, J., Ogino, C., Kondo, A.
    Capsid-like particles consisting of a hepatitis B core (HBc) protein have been studied for their potential as carriers for drug delivery systems (DDS). The hollow HBc particle, which is formed by the self-assembly of core proteins comprising 183 aa residues, has the ability to bind to various cells non-specifically via the action of an arginine-rich domain. In this study, we developed an engineered HBc particle that specifically recognizes and targets human epidermal growth factor receptor-related 2 (HER2)-expressing breast cancer cells. To despoil the non-specific binding property of an HBc particle, we genetically deleted the C-terminal 150-183 aa part of the core protein that encodes the arginine-rich domain (delta HBc). Then, we genetically inserted a Z(HER2) affibody molecule into the 78-81 aa position of the core protein to confer the ability of target-cell-specific recognition. The constructed Z(HER2)-displaying HBc (Z(HER2)-delta HBc) particle specifically recognized HER2-expressing SKBR3 and MCF-7 breast cancer cells. In addition, the Z(HER2)-delta HBc particle exhibited different binding amounts in accordance with the HER2 expression levels of cancer cells. These results show that the display of other types of affibody molecules on HBc particles would be an expandable strategy for targeting several kinds of cancer cells that would help enable a pinpoint DDS.
    OXFORD UNIV PRESS, Mar. 2013, Journal of Biochemistry, 153(3) (3), 251 - 256, English
    [Refereed]
    Scientific journal

  • Nishimura, Y., Takeda, K., Ishii, J., Ogino, C., Kondo, A.
    A bio-nanocapsule derived from the hepatitis B virus (HBV) is expected to be useful as a drug delivery system carrier. Because various types of bio-nanocapsules have been developed, a simple and versatile purification method for bio-nanocapsules would be useful. Therefore, this study was focused on establishing a simple purification method using affinity chromatography by inserting a histidine tag (His-tag) into a bio-nanocapsule. The method achieved a simple, one-step purification with a yield that was 2.5-fold higher than conventional ultracentrifugation, and thus would be a desirable alternative method for recombinant virus-like particle purification. Crown Copyright (c) 2013 Published by Elsevier B.V. All rights reserved.
    ELSEVIER SCIENCE BV, May 2013, Journal of Virological Methods, 189(2) (2), 393 - 396, English
    [Refereed]
    Scientific journal

  • Hasunuma, T., Okazaki, F., Okai, N., Hara, K.Y., Ishii, J., Kondo, A.
    The biorefinery manufacturing process for producing chemicals and liquid fuels from biomass is a promising approach for securing energy and resources. To establish cost-effective fermentation of lignocellulosic biomass, the consolidation of sacccharification and fermentation processes is a desirable strategy, but requires the development of microorganisms capable of cellulose/hemicellulose hydrolysis and target chemical production. Such an endeavor requires a large number of prerequisites to be realized, including engineering microbial strains with high cellulolytic activity, high product yield, productivities, and titers, ability to use many carbon sources, and resistance to toxic compounds released during the pretreatment of lignocellulosic biomass. Researchers have focused on either engineering naturally cellulolytic microorganisms to improve product-related properties or modifying non-cellulolytic organisms with high product yields to become cellulolytic. This article reviews recent advances in the development of microorganisms for the production of renewable chemicals and advanced biofuels, as well as ethanol, from lignocellulosic materials through consolidated bioprocessing. (C) 2012 Elsevier Ltd. All rights reserved.
    ELSEVIER SCI LTD, May 2013, Bioresource Technology, 135, 513 - 522, English
    [Refereed]
    Scientific journal

  • Nishimura, Y., Mieda, H., Ishii, J., Ogino, C., Fujiwara, T., Kondo, A.
    Background: Small interfering RNA (siRNA) has attracted attention in the field of nucleic acid medicine as a RNA interference (RNAi) application that leads to gene silencing due to specific messenger RNA (mRNA) destruction. However, since siRNA is unstable in blood and unable to cross the cell membrane, encapsulation of siRNA into a carrier is required. Results: In this study, we used a carrier that combined Z(HER2)-displaying bio-nanocapsule (derived from hepatitis B virus surface antigen) and liposomes in a complex in order to investigate the feasibility of effective and target-cell-specific RNAi applications. As a result, by observing RNAi only in HER2-expressing breast cancer cells, using our proposed methodology, we successfully demonstrated target-cell-specific delivery and effective function expression of siRNA. Conclusions: These findings show that, in the field of nucleic acid medicine, Z(HER2)-BNC/ LP can be a useful carrier for siRNA delivery, and could also become a useful tool for gene silencing and to accomplish protein knock-down.
    BIOMED CENTRAL LTD, Jun. 2013, Journal of Nanobiotechnology, 11(1) (1), 19, English
    [Refereed]
    Scientific journal

  • Nakamura, Y., Ishii, J., Kondo, A.
    Potentially immeasurable heterodimer combinations of human G-protein-coupled receptors (GPCRs) result in a great deal of physiological diversity and provide a new opportunity for drug discovery. However, due to the existence of numerous combinations, the sets of GPCR dimers are almost entirely unknown and thus their dominant roles are still poorly understood. Thus, the identification of GPCR dimer pairs has been a major challenge. Here, we established a specialized method to screen potential heterodimer partners of human GPCRs based on the split-ubiquitin membrane yeast two-hybrid system. We demonstrate that the mitogen-activated protein kinase (MAPK) signal-independent method can detect ligand-induced conformational changes and rapidly identify heterodimer partners for target GPCRs. Our data present the abilities to apply for the intermolecular mapping of interactions among GPCRs and to uncover potential GPCR targets for the development of new therapeutic agents.
    PUBLIC LIBRARY SCIENCE, Jun. 2013, PLoS ONE, 8(6) (6), e66793, English
    [Refereed]
    Scientific journal

  • Matsuda, F., Shirai, T., Ishii, J., Kondo, A.
    Metabolic inhibitors were applied for chemical regulation of central carbon metabolism in Saccharomyces cerevisiae. S. cerevisiae was treated with 10 metabolic inhibitors with various modes of action, and their activities were evaluated using a growth inhibition assay. Among the 6 active inhibitors, the effects of pyrazole (alcohol dehydrogenase inhibitor) and TTA (2-thenoyltrifluoloacetone, succinate dehydrogenase inhibitor) were analyzed in detail. The flask-scale batch-fermentation test showed that ethanol yield was reduced to 0.10 +/- 0.01 g g(-1) and glycerol yield increased to 0.26 +/- 0.01 g g(-1) on treatment with pyrazole at 5.0 g L-1, indicating that multiple isozymes of alcohol dehydrogenase were simultaneously inhibited. The multi-targeted metabolic profiling analysis revealed that, although the TTA and pyrazole treatments affected the profiles of all central carbon metabolites in distinct manners, the level of fructose-1,6-bisphosphate commonly increased in the TTA- and pyrazole-treated S. cerevisiae by an unknown mechanism. These results demonstrate that chemical regulation of the central carbon metabolism could be used as an alternative tool to control microbial cell factories for bioproduction, or as a chemical probe to investigate the metabolic systems of useful microorganisms. (C) 2013, The Society for Biotechnology, japan. All rights reserved.
    SOC BIOSCIENCE BIOENGINEERING JAPAN, Jul. 2013, Journal of Bioscience and Bioengineering, 116(1) (1), 59 - 64, English
    [Refereed]
    Scientific journal

  • Matsuda, F., Ishii, J., Kondo, T., Ida, K., Tezuka, H., Kondo, A.
    Background: Isobutanol is an important target for biorefinery research as a next-generation biofuel and a building block for commodity chemical production. Metabolically engineered microbial strains to produce isobutanol have been successfully developed by introducing the Ehrlich pathway into bacterial hosts. Isobutanol-producing baker's yeast (Saccharomyces cerevisiae) strains have been developed following the strategy with respect to its advantageous characteristics for cost-effective isobutanol production. However, the isobutanol yields and titers attained by the developed strains need to be further improved through engineering of S. cerevisiae metabolism. Results: Two strategies including eliminating competing pathways and resolving the cofactor imbalance were applied to improve isobutanol production in S. cerevisiae. Isobutanol production levels were increased in strains lacking genes encoding members of the pyruvate dehydrogenase complex such as LPD1, indicating that the pyruvate supply for isobutanol biosynthesis is competing with acetyl-CoA biosynthesis in mitochondria. Isobutanol production was increased by overexpression of enzymes responsible for transhydrogenase-like shunts such as pyruvate carboxylase, malate dehydrogenase, and malic enzyme. The integration of a single gene deletion lpd1 Delta and the activation of the transhydrogenase-like shunt further increased isobutanol levels. In a batch fermentation test at the 50-mL scale from 100 g/L glucose using the two integrated strains, the isobutanol titer reached 1.62 +/- 0.11 g/L and 1.61 +/- 0.03 g/L at 24 h after the start of fermentation, which corresponds to the yield at 0.016 +/- 0.001 g/g glucose consumed and 0.016 +/- 0.0003 g/g glucose consumed, respectively. Conclusions: These results demonstrate that downregulation of competing pathways and metabolic functions for resolving the cofactor imbalance are promising strategies to construct S. cerevisiae strains that effectively produce isobutanol.
    BIOMED CENTRAL LTD, Dec. 2013, Microbial Cell Factories, 12(1) (1), 119, English
    [Refereed]
    Scientific journal

  • Nakamura, Y., Ishii, J., Kondo, A.
    G-protein-coupled receptors (GPCRs) are currently the most important pharmaceutical targets for drug discovery because they regulate a wide variety of physiological processes. Consequently, simple and convenient detection systems for ligands that regulate the function of GPCR have attracted attention as powerful tools for new drug development. We previously developed a yeast-based fluorescence reporter ligand detection system using flow cytometry. However, using this conventional detection system, fluorescence from a cell expressing GFP and responding to a ligand is weak, making detection of these cells by fluorescence microscopy difficult. We here report improvements to the conventional yeast fluorescence reporter assay system resulting in the development of a new highly-sensitive fluorescence reporter assay system with extremely bright fluorescence and high signal-to-noise (S/N) ratio. This new system allowed the easy detection of GPCR signaling in yeast using fluorescence microscopy. Somatostatin receptor and neurotensin receptor (implicated in Alzheimer's disease and Parkinson's disease, respectively) were chosen as human GPCR(s). The facile detection of binding to these receptors by cognate peptide ligands was demonstrated. In addition, we established a highly sensitive ligand detection system using yeast cell surface display technology that is applicable to peptide screening, and demonstrate that the display of various peptide analogs of neurotensin can activate signaling through the neurotensin receptor in yeast cells. Our system could be useful for identifying lead peptides with agonistic activity towards targeted human GPCR(s).
    PUBLIC LIBRARY SCIENCE, Dec. 2013, PLoS ONE, 8(12) (12), e82237, English
    [Refereed]
    Scientific journal

  • Fukutani, Y., Nakamura, T., Yorozu, M., Ishii, J., Kondo, A., Yohda, M.
    For the development of a biomimetic odor-sensing system, we investigated the effects of replacing the N-terminus of an olfactory receptor (OR) on its functional expression in the budding yeast, Saccharomyces cerevisiae. Using the mouse olfactory receptor OR226 (mOR226), three types of chimeric ORs were constructed by replacing N-terminal regions of mOR226 with the corresponding regions of the rat I7 receptor, which is known to be functionally expressed in yeast. The replacement of the N-terminal region of mOR226 dramatically affected the expression and localization of the receptor and improved the sensing ability of the yeast cells for the odorant. Furthermore, the replacement of the endogenous yeast G-protein a subunit (Gpa1) by the OR-specific Golf drastically elevated the odorant-sensing ability of the yeast cells and caused the cells to display a dose-dependent responsiveness to the odorant. Because of the suitability of yeast cells for screening large-scale libraries, the strategy presented here would be useful for the establishment of advanced biomimetic odor-sensing systems. Biotechnol. Bioeng. 2012;109: 205212. (c) 2011 Wiley Periodicals, Inc.
    WILEY-BLACKWELL, Jan. 2012, Biotechnology and Bioengineering, 109(1) (1), 205 - 212, English
    [Refereed]
    Scientific journal

  • Nishimura, Y., Shishido, T., Ishii, J., Tanaka, T., Ogino, C., Kondo, A.
    Bio-nanocapsules (BNCs) are hollow nanoparticles composed of the L protein of hepatitis B virus (HBV) surface antigen (HBsAg), which can specifically introduce genes and drugs into various kinds of target cells. Although the classic electroporation method has typically been used to introduce highly charged molecules such as DNA, it is rarely adopted for proteins due to its very low efficiency. In this study, a novel approach to the preparation of BNC was established whereby a target protein was pre-encapsulated during the course of nanoparticle formation. Briefly, because of the process of BNC formation in a budding manner on the endoplasmic reticulum (ER) membrane, the association of target proteins to the ER membrane with lipidation sequences (ER membrane localization sequences) could directly generate protein-encapsulating BNC in collaboration with co-expression of the L proteins. Since the membrane-localized proteins are automatically enveloped into BNCs during the budding event, this method can be protect the proteins and BNCs from damage caused by electroporation and obviate the need for laborious consideration to study the optimal conditions for protein encapsulation. This approach would be a useful method for encapsulating therapeutic candidate proteins into BNCs. (C) 2011 Elsevier B.V. All rights reserved.
    ELSEVIER SCIENCE BV, Jan. 2012, Journal of Biotechnology, 157(1) (1), 124 - 129, English
    [Refereed]
    Scientific journal

  • Ryo, S., Ishii, J., Iguchi, Y., Fukuda, N., Kondo, A.
    Here we present a successful transplantation of the GAL genetic regulatory circuitry into the G-protein signaling pathway in yeast. The GAL regulon represents a strictly regulated transcriptional mechanism that we have transplanted into yeast to create a highly robust induction system to assist the detection of on-off switching in G-protein signaling. In our system, we engineered yeast to drive the positive GAL regulatory gene in response to agonist-promoted G-protein signaling and to induce transcription of a green fluorescent protein (GFP) reporter gene under the control of the GAL structural gene promoter. Consequently, in response to agonist stimulation of G-protein-coupled receptors (GPCRs), the engineered yeast achieved more than a 150-fold increase in reporter intensity in up to 98% of cells, as determined by flow cytometric sorting. Surprisingly, agonist-stimulated induction of the GFP reporter gene was higher than that by galactose. Our approach to boost reporter gene induction could be applicable in establishing more efficient yeast-based flow cytometric screening systems for agonistic ligands for heterogeneous GPCRs. (c) 2012 Elsevier Inc. All rights reserved.
    ACADEMIC PRESS INC ELSEVIER SCIENCE, May 2012, Analytical Biochemistry, 424(1) (1), 27 - 31, English
    [Refereed]
    Scientific journal

  • Kondo, T., Tezuka, H., Ishii, J., Matsuda, F., Ogino, C., Kondo, A.
    The production of higher alcohols by engineered bacteria has received significant attention. The budding yeast, Saccharomyces cerevisiae, has considerable potential as a producer of higher alcohols because of its capacity to naturally fabricate fusel alcohols, in addition to its robustness and tolerance to low pH. However, because its natural productivity is not significant, we considered a strategy of genetic engineering to increase production of the branched-chain higher alcohol isobutanol, which is involved in valine biosynthesis. Initially, we overexpressed 2-keto acid decarboxylase (KDC) and alcohol dehydrogenase (ADH) in S. cerevisiae to enhance the endogenous activity of the Ehrlich pathway. We then overexpressed Ilv2, which catalyzes the first step in the valine synthetic pathway, and deleted the PDC1 gene encoding a major pyruvate decarboxylase with the intent of altering the abundant ethanol flux via pyruvate. Through these engineering steps, along with modification of culture conditions, the isobutanol titer of S. cerevisiae was elevated 13-fold, from 11 mg/l to 143 mg/l, and the yield was 6.6 mg/g glucose, which is higher than any previously reported value for S. cerevisiae. (C) 2012 Elsevier B.V. All rights reserved.
    ELSEVIER SCIENCE BV, May 2012, Journal of Biotechnology, 159(1-2) (1-2), 32 - 37, English
    [Refereed]
    Scientific journal

  • Ishii, J., Yoshimoto, N., Tatematsu, K., Kuroda, S., Ogino, C., Fukuda, H., Kondo, A.
    G-protein-coupled receptors (GPCRs) regulate a wide variety of physiological processes and are important pharmaceutical targets for drug discovery. Here, we describe a unique concept based on yeast cell-surface display technology to selectively track eligible peptides with agonistic activity for human GPCRs (Cell Wall Trapping of Autocrine Peptides (CWTrAP) strategy). In our strategy, individual recombinant yeast cells are able to report autocrine-positive activity for human GPCRs by expressing a candidate peptide fused to an anchoring motif. Following expression and activation, yeast cells trap autocrine peptides onto their cell walls. Because captured peptides are incapable of diffusion, they have no impact on surrounding yeast cells that express the target human GPCR and non-signaling peptides. Therefore, individual yeast cells can assemble the autonomous signaling complex and allow single-cell screening of a yeast population. Our strategy may be applied to identify eligible peptides with agonistic activity for target human GPCRs.
    PUBLIC LIBRARY SCIENCE, May 2012, PLoS ONE, 7(5) (5), e37136, English
    [Refereed]
    Scientific journal

  • Hara, K.Y., Kim, S., Kiriyama, K., Yoshida, H., Arai, S., Ishii, J., Ogino, C., Fukuda, H., Kondo, A.
    Glutathione is a valuable tripeptide that is widely used in the pharmaceutical, food, and cosmetic industries. Glutathione is industrially produced by fermentation using Saccharomyces cerevisiae. Before the glutathione fermentation process with S. cerevisiae, a glucose extraction process from starchy materials is required. This glucose extraction is usually carried out by converting starchy materials to starch using high-temperature cooking and subsequent hydrolysis by amylases to convert starch to glucose. In this study, to develop an energy-saving glutathione production process by reducing energy consumption during the cooking step, we efficiently produced glutathione from low-temperature cooked rice using amylase-expressing S. cerevisiae. The combination of the amylase-expressing yeast with low-temperature cooking is potentially applicable to a variety of energy-saving bio-production methods of chemicals from starchy bio-resources.
    WILEY-BLACKWELL, May 2012, Biotechnology Journal, 7(5) (5), 686 - 689, English
    [Refereed]
    Scientific journal

  • Ishii, J., Moriguchi, M., Hara, K.Y., Shibasaki, S., Fukuda, H., Kondo, A.
    Flow cytometry enables comparative quantification, population analysis, and high-throughput screening of agonist-mediated G-protein-coupled receptor (GPCR) signaling in genetically engineered yeasts. By using flow cytometry, we found that transformation of yeast cells with a low plasmid number is critical both for the construction of large screening libraries and for stable signal transmission in cell ensembles. Based on these findings, we constructed an engineered yeast strain for the improved identification of signal promotion by C alpha(i)-specific human GPCRs using flow cytometry. (C) 2012 Elsevier Inc. All rights reserved.
    ACADEMIC PRESS INC ELSEVIER SCIENCE, Jul. 2012, Analytical Biochemistry, 426(2) (2), 129 - 133, English
    [Refereed]
    Scientific journal

  • Matsuda, F., Kondo, T., Ida, K., Tezuka, H., Ishii, J., Kondo, A.
    To increase isobutanol production in Saccharomyces cerevisiae, the valine biosynthetic pathway was activated by overexpression of the relevant enzymes in the mitochondria and the cytosol. Native mitochondrial enzymes were overepxressed in the cytosol by deleting the mitochondrial transit peptides. The metabolically engineered S. cerevisiae possessing the cytosolic pathway showed increased isobutanol production (63 +/- 4 mg/L).
    TAYLOR & FRANCIS LTD, Nov. 2012, Bioscience, Biotechnology and Biochemistry, 76(11) (11), 2139 - 2141, English
    [Refereed]
    Scientific journal

  • Nishimura, Y., Ishii, J., Okazaki, F., Ogino, C., Kondo, A.
    A bio-nanocapsule (BNC), a hollow particle composed of hepatitis B virus (HBV) surface antigen (HBsAg), and liposome (LP) conjugation method (BNC/LP) has been recently developed by Jung et al. (2008). The BNC/LP complex carrier could successfully deliver fluorescence-labeled beads (100 nm) into liver cells. In this study, we report the promising delivery of proteins incorporated in the complex carriers, which were prepared by the BNC/LP conjugation method with specificity-altered BNC and composition-varied LPs. The specificity-altered BNC, Z(HER2)-BNC was developed by replacing the hepatocyte recognition site of BNC with Z(HER2) binding to HER2 receptor specifically. Using green fluorescent protein (GFP; 27 kDa) and cellular cytotoxic protein (exotoxin A; 66 kDa) for the delivery, we herein present the impact of different charges attributed to the composition of the LP on specific cell targeting and cellular uptake of the complex carriers. In addition, we demonstrate that the mixture prepared by mixing LPs with helper lipid possessing endosomal escaping ability boosts the functional expression of the cellular cytotoxic exotoxin A activity specifically. Finally, we further show the blending ratio of the LP mixture and Z(HER2)-BNC is a critical factor in determining the highly-efficient expression of the cytotoxic activity of exotoxin A.
    INFORMA HEALTHCARE, Dec. 2012, Journal of Drug Targeting, 20(10) (10), 897 - 905, English
    [Refereed]
    Scientific journal

  • Fukutani, Y., Ishii, J., Noguchi, K., Kondo, A., Yohda, M.
    The goal of this work was to improve the bioluminescence-based signaling assay system to create a practical application of a biomimetic odor sensor using an engineered yeast-expressing olfactory receptors (ORs). Using the yeast endogenous pheromone receptor (Ste2p) as a model GPCR, we determined the suitable promoters for the firefly luciferase (luc) reporter and GPCR genes. Additionally, we deleted some genes to further improve the sensitivity of the luc reporter assay. By replacing the endogenous yeast G-protein a-subunit (Gpa1p) with the olfactory-specific Gaolf, the optimized yeast strain successfully transduced signal through both OR and yeast Ste2p. Our results will assist the development of a bioluminescence-based odor-sensing system using OR-expressing yeast. Biotechnol. Bioeng. 2012; 109: 31433151. (C) 2012 Wiley Periodicals, Inc.
    WILEY, Dec. 2012, Biotechnology and Bioengineering, 109(12) (12), 3143 - 3151, English
    [Refereed]
    Scientific journal

  • Hasunuma, T., Sanda, T., Yamada, R., Yoshimura, K., Ishii, J., Kondo, A.
    Background: The development of novel yeast strains with increased tolerance toward inhibitors in lignocellulosic hydrolysates is highly desirable for the production of bio-ethanol. Weak organic acids such as acetic and formic acids are necessarily released during the pretreatment (i.e. solubilization and hydrolysis) of lignocelluloses, which negatively affect microbial growth and ethanol production. However, since the mode of toxicity is complicated, genetic engineering strategies addressing yeast tolerance to weak organic acids have been rare. Thus, enhanced basic research is expected to identify target genes for improved weak acid tolerance. Results: In this study, the effect of acetic acid on xylose fermentation was analyzed by examining metabolite profiles in a recombinant xylose-fermenting strain of Saccharomyces cerevisiae. Metabolome analysis revealed that metabolites involved in the non-oxidative pentose phosphate pathway (PPP) [e.g. sedoheptulose-7-phosphate, ribulose-5-phosphate, ribose-5-phosphate and erythrose-4-phosphate] were significantly accumulated by the addition of acetate, indicating the possibility that acetic acid slows down the flux of the pathway. Accordingly, a gene encoding a PPP-related enzyme, transaldolase or transketolase, was overexpressed in the xylose-fermenting yeast, which successfully conferred increased ethanol productivity in the presence of acetic and formic acid. Conclusions: Our metabolomic approach revealed one of the molecular events underlying the response to acetic acid and focuses attention on the non-oxidative PPP as a target for metabolic engineering. An important challenge for metabolic engineering is identification of gene targets that have material importance. This study has demonstrated that metabolomics is a powerful tool to develop rational strategies to confer tolerance to stress through genetic engineering.
    BIOMED CENTRAL LTD, Jan. 2011, Microbial Cell Factories, 10, English
    [Refereed]
    Scientific journal

  • Fukuda, N., Ishii, J., Kondo, A.
    Weak and transient protein-protein interactions are associated with biological processes, but many are still undefined because of the difficulties in their identification. Here, we describe a redesigned method to screen transient protein-protein interactions by using a novel signal amplification circuit, which is incorporated into yeast to artificially magnify the signal responding to the interactions. This refined method is based on the previously established G gamma recruitment system, which utilizes yeast G-protein signaling and mating growth selection to screen interacting protein pairs. In the current study, to test the capability of our method, we chose mutants of the Z-domain derived from Staphylococcus aureus protein A as candidate proteins, and the Fc region of human IgG as the counterpart. By introduction of an artificial signal amplifier into the previous G gamma recruitment system, the signal transduction responding to transient interactions between Z-domain mutants and the Fc region with significantly low affinity (8.0 x 10(3) M(-1)) was successfully amplified in recombinant haploid yeast cells. As a result of zygosis with the opposite mating type of wildtype haploid cells, diploid colonies were vigorously and selectively generated on the screening plates, whereas our previous system rarely produced positive colonies. This new approach will be useful for exploring the numerous transient interactions that remain undefined because of the lack of powerful screening tools for their identification.
    WILEY-BLACKWELL, Sep. 2011, FEBS Journal, 278(17) (17), 3086 - 3094, English
    [Refereed]
    Scientific journal

  • Matsuda, F., Furusawa, C., Kondo, T., Ishii, J., Shimizu, H., Kondo, A.
    Background: While Saccharomyces cerevisiae is a promising host for cost-effective biorefinary processes due to its tolerance to various stresses during fermentation, the metabolically engineered S. cerevisiae strains exhibited rather limited production of higher alcohols than that of Escherichia coli. Since the structure of the central metabolism of S. cerevisiae is distinct from that of E. coli, there might be a problem in the structure of the central metabolism of S. cerevisiae. In this study, the potential production of higher alcohols by S. cerevisiae is compared to that of E. coli by employing metabolic simulation techniques. Based on the simulation results, novel metabolic engineering strategies for improving higher alcohol production by S. cerevisiae were investigated by in silico modifications of the metabolic models of S. cerevisiae. Results: The metabolic simulations confirmed that the high production of butanols and propanols by the metabolically engineered E. coli strains is derived from the flexible behavior of their central metabolism. Reducing this flexibility by gene deletion is an effective strategy to restrict the metabolic states for producing target alcohols. In contrast, the lower yield using S. cerevisiae originates from the structurally limited flexibility of its central metabolism in which gene deletions severely reduced cell growth. Conclusions: The metabolic simulation demonstrated that the poor productivity of S. cerevisiae was improved by the introduction of E. coli genes to compensate the structural difference. This suggested that gene supplementation is a promising strategy for the metabolic engineering of S. cerevisiae to produce higher alcohols which should be the next challenge for the synthetic bioengineering of S. cerevisiae for the efficient production of higher alcohols.
    BIOMED CENTRAL LTD, Sep. 2011, Microbial Cell Factories, 10, English
    [Refereed]
    Scientific journal

  • Fukuda, N., Ishii, J., Kaishima, M., Kondo, A.
    G-protein-coupled receptors (GPCRs) are considered as important targets for drug discovery. The yeast Saccharomyces cerevisiae is an attractive host for high-throughput screening of agonistic ligands for human GPCRs because it can simplify the complicated signaling pathways that are present in mammalian cell lines. Unfortunately, many human GPCRs induce only partial signal activation in yeast cells depending on their coupling efficiency with yeast G-proteins. This problem often results in unsatisfactory detection sensitivity, thereby resulting in a limitation to yeast-based detection systems. Here we introduce a new highly sensitive detection method that provides robust agonist detection of human GPCRs. Our strategy is designed to invoke feedback activation of signals within yeast G-protein signaling pathways. Briefly, agonist stimulation of human GPCRs triggers expression of an artificial signal activator that amplifies signaling. We chose human somatostatin receptor subtype 5 (hSSTR5) as a model of a human GPCR. Investigation of the response of hSSTR5-expressing yeast to various concentrations of somatostatin demonstrated that feedback activation of the signal can successfully improve the detection limit and the maximum level of signaling. This novel approach will enhance the usefulness of yeast-based screening of agonistic ligands for a variety of human GPCRs. (C) 2011 Elsevier Inc. All rights reserved.
    ACADEMIC PRESS INC ELSEVIER SCIENCE, Oct. 2011, Analytical Biochemistry, 417(2) (2), 182 - 187, English
    [Refereed]
    Scientific journal

  • Tanino, T., Hotta, A., Ito, T., Ishii, J., Yamada, R., Hasunuma, T., Ogino, C., Ohmura, N., Ohshima, T., Kondo, A.
    A yeast with the xylose isomerase (XI) pathway was constructed by the multicopy integration of XI overexpression cassettes into the genome of the Saccharomyces cerevisiae MT8-1 strain. The resulting yeast strain successfully produced ethanol from both xylose as the sole carbon source and a mixed sugar, consisting of xylose and glucose, without any adaptation procedure. Ethanol yields in the fermentation from xylose and mixed sugar were 61.9% and 62.2% of the theoretical carbon recovery, respectively. Knockout of GRE3, a gene encoding nonspecific aldose reductase, of the host yeast strain improved the fermentation profile. Not only specific ethanol production rates but also xylose consumption rates was improved more than twice that of xylose-metabolizing yeast with the XI pathway using GRE3 active yeast as the host strain. In addition, it was demonstrated that xylitol in the medium exhibits a concentration-dependent inhibition effect on the ethanol production from xylose with the yeast harboring the XI-based xylose metabolic pathway. From our findings, the combination of XI-pathway integration and GRE3 knockout could be result in a consolidated xylose assimilation pathway and increased ethanol productivity.
    SPRINGER, Nov. 2010, Applied Microbiology and Biotechnology, 88(5) (5), 1215 - 1221, English
    [Refereed]
    Scientific journal

  • 野坂 和輝, 宮地 佑典, 石井 純, 荻野 千秋, 近藤 昭彦
    公益社団法人 化学工学会, 2010, 化学工学会 研究発表講演要旨集, 2010, 983 - 983, Japanese

  • 漁 慎太郎, 井口 裕介, 福田 展雄, 石井 純, 近藤 昭彦
    公益社団法人 化学工学会, 2010, 化学工学会 研究発表講演要旨集, 2010, 977 - 977, Japanese

  • Tanaka, T., Masunari, S., Ishii, J., Wakamura, K., Segawa, M., Fukuda, H., Kondo, A.
    Here we expand the yeast cell surface display system to display non-natural, functional molecules. The short biotin acceptor peptide (BAP) sequence of biotin ligase from E. coli(BirA) was genetically introduced to the N-terminus of the anchor protein, Flo428. Through co-expression of BAP-fused Flo428 with BirA, biotinylated BAP could be displayed on the yeast cell surface. Subsequent addition of streptavidin-FITC resulted in the display of streptavidin-FITC, and, the display of biotin-FITC was successful using streptavidin as a linker. Our strategy provides a powerful tool for displaying functional molecules on yeast cell surfaces. (c) 2009 Elsevier B.V. All rights reserved.
    ELSEVIER SCIENCE BV, Jan. 2010, Journal of Biotechnology, 145(1) (1), 79 - 83, English
    [Refereed]
    Scientific journal

  • Fukuda, N., Ishii, J., Tanaka, T., Kondo, A.
    We have developed a new approach based on the G gamma recruitment system to screen affinity-enhanced proteins by expressing a binding competitor. The previously established G gamma recruitment system is a yeast two-hybrid (Y2H) system that utilizes G-protein signaling, and is based on the fact that membrane localization of the G-protein c subunit (G gamma) is essential for signal transduction in yeast. In the original Y2H system, an engineered G gamma that lacks membrane localization upon deletion of the lipid modi. cation site (G gamma(cyto)) is produced, and a candidate protein with an artificial lipidation site and its counterpart fused with G gamma(cyto) are expressed. As protein-protein interactions bring G gamma(cyto) towards the plasma membrane, G-protein signaling can be activated, and the interaction is detected by various cellular responses as the readout. In the current study, we expressed a third cytosolic protein that competes with the candidate protein to specifically isolate affinity-enhanced mutants from a mutation library of the candidate protein. Enhancing the affinity of the protein candidate guides the counterpart-G gamma(cyto) fusion protein towards the plasma membrane and activates signaling. Using mutants of the Z domain derived from Staphylococcus aureus protein A as candidate proteins or competitors, and the Fc portion of human immunoglobulin G (IgG) as the counterpart, we demonstrate that affinity-enhanced proteins can be effectively screened from a library containing a 10 000-fold excess of non-enhanced proteins. This new approach, called the competitor-introduced G gamma recruitment system, will be useful for efficient discovery of rare valuable candidates hidden among excess ordinary ones.
    WILEY-BLACKWELL PUBLISHING, INC, Apr. 2010, FEBS Journal, 277(7) (7), 1704 - 1712, English
    [Refereed]
    Scientific journal

  • Togawa, S., Ishii, J., Ishikura, A., Tanaka, T., Ogino, C., Kondo, A.
    N-linked oligosaccharides or asparagine residues are often involved in G protein-coupled receptor functions. Focusing on Asn13 and Asn26 positioned on N-linked glycosylation motifs in the amino-terminal domain of human somatostatin receptor subtype-5 (hSSTR5), we performed site-directed mutagenesis and evaluated the mutants by using yeast cells as the host strain. This is because analysing the complicated signalling in mammalian cell lines is simplified by the utilization of the monopolistic pheromone signalling pathway in yeast. Western blot analysis and confocal laser scanning microscope observation showed that Asn13 and/or Asn26 mutations had no effects on cell-surface expression of hSSTR5 in yeast. By using an engineered yeast strain of Saccharomyces cerevisiae, which induces the expression of the green fluorescent protein (GFP) reporter gene in response to the agonist-specific signal transduction, it was demonstrated that a single mutation of two asparagine residues attenuated the somatostatin-specific signalling levels, and the double mutant significantly lost the signalling ability. These results clearly show the importance of these asparagine residues in the agonist-specific signalling of hSSTR5, although it was not enough to identify the consequence of oligosaccharides.
    OXFORD UNIV PRESS, Jun. 2010, Journal of Biochemistry, 147(6) (6), 867 - 873, English
    [Refereed]
    Scientific journal

  • Iguchi, Y., Ishii, J., Nakayama, H., Ishikura, A., Izawa, K., Tanaka, T., Ogino, C., Kondo, A.
    The yeast Saccharomyces cerevisiae is known as an available host for human G-protein-coupled receptor (GPCR) ligand screening. Although several types of yeast signal sequences (SS) attached with the GPCRs could improve their productivities and facilitate transportation of the GPCRs to the yeast plasma membrane, the effects of additional SS on ligand-specific signalling functions of GPCRs are not reported. Here, we demonstrated the controlling signalling properties by addition of SS using engineered yeast as a host. Prepro and pre regions of alpha-factor and amino-terminal sequence of Ste2 (Ste2N) were used as SS, and somatostatin (SST) receptor subtype-5 (SSTR5) was used as a model GPCR. We also constructed a yeast-based fluorescent assay system for monitoring the activation levels of SSTR5 signalling by a green fluorescent protein (GFP) reporter gene. The production levels and localisation patterns of the SS-attached SSTR5 were more significantly improved than those of wild-type SSTR5. In addition, we successfully controlled the pharmacological efficacy and potency by introducing SS. Among four types of SSTR5 receptors, Ste2N-SSTR5 responded at the lowest ligand concentration. This finding will be informative for constructing optimal yeast-based ligand screening systems to discriminate the cells on the basis of signalling levels.
    OXFORD UNIV PRESS, Jun. 2010, Journal of Biochemistry, 147(6) (6), 875 - 884, English
    [Refereed]
    Scientific journal

  • Ishii, J., Fukuda, N., Tanaka, T., Ogino, C., Kondo, A.
    For elucidating protein-protein interactions, many methodologies have been developed during the past two decades. For investigation of interactions inside cells under physiological conditions, yeast is an attractive organism with which to quickly screen for hopeful candidates using versatile genetic technologies, and various types of approaches are now available. Among them, a variety of unique systems using the guanine nucleotide-binding protein (G-protein) signaling pathway in yeast have been established to investigate the interactions of proteins for biological study and pharmaceutical research. G-proteins involved in various cellular processes are mainly divided into two groups: small monomeric G-proteins, and heterotrimeric G-proteins. In this minireview, we summarize the basic principles and applications of yeast-based screening systems, using these two types of G-protein, which are typically used for elucidating biological protein interactions but are differentiated from traditional yeast two-hybrid systems.
    Lead, WILEY-BLACKWELL, May 2010, FEBS Journal, 277(9) (9), 1982 - 1995, English, International magazine
    [Refereed]
    Scientific journal

  • CONSTRUCTION OF A NOVEL DETECTION SYSTEM FOR PROTEIN-PROTEIN INTERACTIONS USING YEAST G-PROTEIN SIGNALING
    Nobuo Fukuda, Jun Ishii, Tsutomu Tanaka, Hideki Fukuda, Akihiko Kondo
    SOUTHEAST UNIV PRESS, 2009, IFPT'6: PROGRESS ON POST-GENOME TECHNOLOGIES, PROCEEDINGS, 238 - 238, English
    [Refereed]
    International conference proceedings

  • Fukuda, N., Ishii, J., Tanaka, T., Fukuda, H., Kondo, A.
    In the current study, we report the construction of a novel system for the detection of protein-protein interactions using yeast G-protein signaling. It is well established that the G-protein gamma subunit (G gamma) is anchored to the inner leaflet of the plasma membrane via lipid modification in the C-terminus, and that this localization of G gamma is required for signal transduction. In our system, mutated G gamma (G gamma(cyto)) lacking membrane localization ability was genetically prepared by deletion of the lipid modification site. Complete disappearance of G-protein signal was observed when G gamma(cyto) was expressed in the cytoplasm of yeast cells from which the endogenous G gamma gene had been deleted. In order to demonstrate the potential use of our system, we utilized the Staphylococcus aureus ZZ domain and the Fc portion of human immunoglobulin G (IgG) as a model interaction pair. To design our detection system for protein-protein interaction, the ZZ domain was altered so that it associates with the inner leaflet of the plasma membrane, and the Fc part was then fused to G gamma(cyto). The Fc-G gamma(cyto) fusion protein migrated towards the membrane via the ZZ-Fc interaction, and signal transduction was therefore restored. This signal was successfully detected by assessing growth inhibition and transcription in response to G-protein signaling. Finally, several Z variants displaying affinity constants ranging from 8.0 x 10(3) to 6.8 x 10(8) m(-1) were prepared, and it was demonstrated that our system was able to discriminate subtle differences in affinity. In conclusion, our system appears to be a reliable and versatile technique for detection of protein-protein interactions, and may prove useful in future protein interaction studies.
    WILEY-BLACKWELL PUBLISHING, INC, May 2009, FEBS Journal, 276(9) (9), 2636 - 2644, English
    [Refereed]
    Scientific journal

  • Ishii, J., Izawa, K., Matsumura, S., Wakamura, K., Tanino, T., Tanaka, T., Ogino, C., Fukuda, H., Kondo, A.
    To allow the comprehensive assessments of yeast expression systems, a simple and immediate method for simultaneously evaluating the expression level and plasmid maintenance in yeast was demonstrated. This method uses green fluorescent protein (GFP) and flow cytometry (FCM) and is characterized by a dual analysis of the average intensity of GFP fluorescence and the population of GFP-expressing cells. The FCM analysis of GFP fluorescence intensity rapidly quantifies the expression level without complex manipulations, such as the enzymatic reaction of a lacZ reporter assay. Moreover, the single-cell analysis revealed that the proportion of cells expressing GFP in the cell cluster reflects the plasmid retention rate; therefore, the FCM analysis of the GFP-expressing population allows the immediate estimation of the plasmid retention rate without the 2- or 3-day incubation required for colony counting. We show that the FCM analysis with GFP reporter is a suitable method to explore the hopeful expression vector and host strain or establish the several expression systems exhibiting the characteristic properties in yeast.
    OXFORD UNIV PRESS, Jun. 2009, Journal of Biochemistry, 145(6) (6), 701 - 708, English
    [Refereed]
    Scientific journal

  • Jun Ishii
    Apr. 2008, Biotechnology Progress, 24(2) (2), English
    [Refereed]
    Scientific journal

  • 若村 香菜子, 田中 勉, 石井 純, 福田 秀樹, 近藤 昭彦
    公益社団法人 化学工学会, 2008, 化学工学会 研究発表講演要旨集, 2008, 944 - 944, Japanese

  • 戸川 翔太, 石井 純, 田中 勉, 立松 健司, 黒田 俊一, 福田 秀樹, 近藤 昭彦
    公益社団法人 化学工学会, 2008, 化学工学会 研究発表講演要旨集, 2008, 971 - 971, Japanese

  • 井口 裕介, 石井 純, 田中 勉, 立松 健司, 黒田 俊一, 福田 秀樹, 近藤 昭彦
    公益社団法人 化学工学会, 2008, 化学工学会 研究発表講演要旨集, 2008, 970 - 970, Japanese

  • 石井 純, 田中 勉, 松村 静香, 立松 健司, 黒田 俊一, 荻野 千秋, 福田 秀樹, 近藤 昭彦
    公益社団法人 化学工学会, 2008, 化学工学会 研究発表講演要旨集, 2008, 348 - 348, Japanese

  • Ishii, J., Tanaka, T., Matsumura, S., Tatematsu, K., Kuroda, S., Ogino, C., Fukuda, H., Kondo, A.
    Here, we describe a yeast-based fluorescence reporter assay for G protein-coupled receptor (GPCR) signalling using a flow cytometer (FCM). The enhanced green fluorescent protein (EGFP) gene was integrated into the FUS1 locus as a reporter gene. The engineered yeast was able to express the EGFP in response to ligand stimulation. Gene-disrupted yeast strains were constructed to evaluate the suitability of the yeast-based fluorescence screening system for heterologous GPCR. When receptor was expressed by episomal plasmid, the proportion of the signalling-activated cells in response to ligand stimulation decreased significantly. The GPCR-signalling-activated and non-activated cell clusters were individually isolated by analysing the fluorescence intensity at the single-cell level with FCM, and it was found that the plasmid retention rate decays markedly in the non-activated cell cluster. We attributed the loss of plasmid to G1 arrest in response to signalling, and successfully improved the plasmid retention rate by disrupting the FAR1 gene and avoiding cell cycle arrest. Our system will be a powerful tool for the quantitative and high-throughput GPCR screening of yeast-based combinatorial libraries using FCM.
    OXFORD UNIV PRESS, May 2008, Journal of Biochemistry, 143(5) (5), 667 - 674, English
    [Refereed]
    Scientific journal

  • Shibasaki, S., Sakata, K., Ishii, J., Kondo, A., Ueda, M.
    A yeast protein fragment complementation assay (PCA) system based on dihydrofolate reductase (DHFR) is difficult to be operated because it is not as sensitive to trimethoprim (TMP) as the system using a prokaryotic microorganism. Here, the PCA system using DHFR, specific inhibitors, and a substrate in the yeast Saccharomyces cerevisiae was newly developed. As a model, the human oncoprotein Ras and the Ras-binding domain (RBD) of Raf-1 were individually and genetically fused to DHFR fragment, and each genetic construct was coexpressed under the control of the GAL1 promoter. An interaction between Ras and RBD could be evaluated on the basis of cell proliferation. To establish the experimental conditions for the yeast PCA system based on the DHFR reconstitution, we examined yeast host strains and the concentration of inhibitory additives to prevent endogenous DHFR activity, namely, TMP and sulfanilamide, and the substrate of DHFR, namely, folic acid. The transformant harboring wild-type Ras or its variants showed positive interaction signals, and the order of interactions for combination corresponded to the results of other in vitro assays. Moreover, combinatorial mutated Ras-binding domains were constructed, and the interaction of RBDs with Ras using this yeast PCA system was examined. As a result, various types of mutated clone for RBD were obtained. These demonstrations suggest that the yeast PCA system based on DHFR can be one of good, convenient, and inexpensive tools for investigating eukaryotic protein-protein interactions in vivo.
    SPRINGER, Sep. 2008, Applied Microbiology and Biotechnology, 80(4) (4), 735 - 743, English
    [Refereed]
    Scientific journal

  • 森口 みゆき, 石井 純, 立松 健司, 黒田 俊一, 福田 秀樹, 近藤 昭彦
    公益社団法人 化学工学会, 2007, 化学工学会 研究発表講演要旨集, 2007, 844 - 844, Japanese

  • 福田 展雄, 石井 純, 芝崎 誠司, 福田 秀樹, 近藤 昭彦
    公益社団法人 化学工学会, 2007, 化学工学会 研究発表講演要旨集, 2007, 325 - 325, Japanese

  • Fluorescence detection system for human G protein - Coupled receptor signaring in yeast
    Jun Ishii, Akihiko Kondo
    PHOENIX PUBL & MEDIA NETWORK, 2007, PROGRESS ON POST-GENOME TECHNOLOGIES, 57 - 57, English
    [Refereed]
    International conference proceedings

  • Shibasaki, S., Kawabata, A., Ishii, J., Yagi, S., Kadonosono, T., Kato, M., Fukuda, N., Kondo, A., Ueda, M.
    We determined whether the cocultivation of yeast cells displaying a ZZ-domain and secreting an Fc fusion protein can be a novel tool for the recovery of secreted recombinant proteins. The ZZ-domain from Staphylococcus aureus protein A was displayed on the cell surface of Saccharomyces cerevisiae under the control of the GAL1 promoter. Strain S. cerevisiae BY4742 cells displaying the ZZ-domain on their surface were used for cocultivation with cells that produce a target protein fused to the Fc fragment as an affinity tag. The enhanced green fluorescent protein or Rhizopus oryzae lipase was genetically fused to the N and C termini of the Fc fragment of human immunoglobulin G, respectively. Through analysis by fluorescence-activated cell sorting and enzymatic assay, it was demonstrated that these fusion proteins are successfully produced in the medium and recovered by affinity binding with the cell surface displaying the ZZ-domain. These results suggest that the ZZ-domain-displaying cell and Fc fusion protein-secreting cell can be applied to use in synergistic process of production and recovery of secreted recombinant proteins.
    SPRINGER, Jun. 2007, Applied Microbiology and Biotechnology, 75(4) (4), 821 - 828, English
    [Refereed]
    Scientific journal

  • Fukuda, N., Ishii, J., Shibasaki, S., Ueda, M., Fukuda, H., Kondo, A.
    We constructed a high-throughput screening (HTS) system for target cells based on the detection of protein-protein interactions by flow cytometric sorting due to the improvement in the yeast cell surface display system. Interaction model proteins, which are the ZZ domain derived from Staphylococcus aureus and the Fe part of human immumoglobulin G (IgG), were displayed on the yeast cell surface. We achieved a rapid and enhanced expression of these proteins as a result of adopting an appropriate yeast strain and a suitable promoter. The displayed ZZ domain had an ability to bind to rabbit IgG and the displayed Fc part to protein A. These were confirmed by flow cytometry and fluorescence microscopy. Furthermore, the cells displaying the ZZ domain or Fc part were isolated from the model libraries constructed by mixing the control yeast cells with the target yeast cells. The ratio of the target cells was increased from 0.0001% to more than 70% by two cycles of cell sorting. These results indicate that we can achieve a rapid and highly efficient isolation method for the target cells with FACSCalibur and that this method will further extend the application of flow cytometric sorting to library selections.
    SPRINGER, Aug. 2007, Applied Microbiology and Biotechnology, 76(1) (1), 151 - 158, English
    [Refereed]
    Scientific journal

  • 森口 みゆき, 石井 純, 立松 健司, 黒田 俊一, 福田 秀樹, 近藤 昭彦
    公益社団法人 化学工学会, 2006, 化学工学会 研究発表講演要旨集, 2006, 986 - 986, Japanese

  • Development of a single cell analysis system of agonist for drug discovery
    Akihiko Kondo, Jun Ishii, Shizuka Matsumura, Hideki Fukuda
    SOUTHEAST UNIV PRESS, 2006, Progress on Post-Genome Technologies, 30 - 30, English
    [Refereed]
    International conference proceedings

  • Ishii, J., Matsumura, S., Kimura, S., Tatematsu, K., Kuroda, S., Fukuda, H., Kondo, A.
    The mechanism of G protein-coupled receptor (GPCR) signaling in yeasts is similar to that in mammalian cells. Therefore, yeasts can be used in GPCR assays, and several ligand detection systems using a pheromone signaling pathway in yeasts have been developed by employing yeasts with disrupted chromosomal genes that code for proteins producing specific effects. In this study, the construction of yeast strains that can detect ligand binding mediated by interactions between the G protein and GPCR using either fluorescence or auxotrophic selectivity is demonstrated. The strain was constructed by integrating the fusion gene of pheromone-responsive protein (FUS1), enhanced green fluorescence protein (EGFP), and auxotrophic marker protein (HIS3) into the FUS1 locus. Moreover, the influence of gene disruptions on the yeast signal transduction cascade is closely investigated with respect to both quantitative and dynamic aspects to further develop a high-throughput screening system for the GPCR assay using yeasts. Yeast strains with a disrupted SST2 gene, which is a member of the RGS (regulator of G protein signaling) family, and a disrupted FAR1 gene, which mediates cell cycle arrest in response to a pheromone, were monitored by measuring their fluorescence and growth rate. This method will be applicable to other comprehensive GPCR ligand screening methods. © 2006 American Chemical Society and American Institute of Chemical Engineers.
    Jul. 2006, Biotechnology Progress, 22(4) (4), 954 - 960, English
    [Refereed]
    Scientific journal

MISC

  • 酵母における遺伝子スイッチ進化工学ワークフローの「ロバスト化」—Robust Evolutionary Engineering Platform for Yeast Genetic Switches
    冨永 将大, 近藤 昭彦, 石井 純
    [京都] : 日本生物物理学会, Jun. 2024, 生物物理 / 日本生物物理学会 編, 64(3) (3), 144 - 146, Japanese

  • ロドプシンによる光駆動プロトンポンプが大腸菌の弱酸耐性に及ぼす影響
    小林祐摩, 佐野海瑚人, 弘埜陽子, 松田史生, 石井純, 原清敬, 戸谷吉博, 清水浩
    2022, 化学工学会年会研究発表講演要旨集(CD-ROM), 87th

  • ロドプシンを利用した光駆動ATP再生によるイソプレノールおよび3ヒドロキシプロピオン酸生産の向上
    戸谷吉博, 佐野海瑚人, 田中涼, 鎌田健太郎, 松田史生, 石井純, 弘埜陽子, 原清敬, 清水浩
    2022, 化学工学会年会研究発表講演要旨集(CD-ROM), 87th

  • Bacterial Rhodopsin reconstruction for improving bioproduction
    佐藤良樹, 川崎真由, 弘埜陽子, 菊川寛史, 菊川寛史, 中野祥吾, 石井純, 松田史生, 戸谷吉博, 伊藤創平, 原清敬, 原清敬
    2022, 日本農芸化学会大会講演要旨集(Web), 2022

  • Improvement of bioalcohol production in Saccharomyces cerevisiae by ATP wasting
    谷田部楓太, 岡橋伸幸, 清家泰介, 石井純, 松田史生
    2022, 日本生物工学会大会講演要旨集, 74th

  • Effect of light-induced proton pump activity by rhodopsin on weak acid resistance in Escherichia coli.
    小林祐摩, 佐野海瑚人, 弘埜陽子, 松田史生, 石井純, 原清敬, 戸谷吉博, 清水浩
    2022, 日本生物工学会大会講演要旨集, 74th

  • Light-driven metabolic conversion from mevalonate to isoprenol in Escherichia coli
    佐野海瑚人, 田中涼, 鎌田健太郎, 松田史生, 石井純, 弘埜陽子, 原清敬, 戸谷吉博, 清水浩
    2021, 日本生物工学会大会講演要旨集, 73rd

  • Bacterial Rhodopsin reconstruction for improving proton pump activity
    佐藤良樹, 川崎真由, 弘埜陽子, 菊川寛史, 中野祥吾, 石井純, 松田史生, 戸谷吉博, 伊藤創平, 原清敬
    2021, 日本生物工学会大会講演要旨集, 73rd

  • Retinal production by engineered Xanthophyllomyces dendrorhous
    藤野紗代, 景山裕也, 弘埜陽子, 菊川寛史, 菊川寛史, 松田史生, 戸谷吉博, 石井純, 原清敬, 原清敬
    2021, 日本生物工学会大会講演要旨集, 73rd

  • Metabolic analysis of mevalonic acid-producing Escherichia coli expressing a light-driven proton pump
    松山千夏, 清家泰介, 岡橋伸幸, 戸谷吉博, 弘埜陽子, 平山英伸, 石井純, 清水浩, 原清敬, 松田史生
    2021, 日本生物工学会大会講演要旨集, 73rd

  • Analysis of deltarhodopsin mutant
    佐藤良樹, 弘埜陽子, 石井純, 柘植謙爾, 松田史生, 戸谷吉博, 原清敬
    2020, 日本農芸化学会大会講演要旨集(Web), 2020

  • Improvement of bioproduction with ATP recycling by delta-rhodopsin
    塩澤優稀, 弘埜陽子, 石井純, 柘植謙爾, 松田史生, 戸谷吉博, 原清敬
    2020, 日本農芸化学会大会講演要旨集(Web), 2020

  • 光駆動ATP再生系を導入したVmax細胞の開発
    弘埜陽子, 戸谷吉博, 石井純, 柘植謙爾, 松田史生, 原清敬
    2019, 日本農芸化学会大会講演要旨集(Web), 2019

  • デルタロドプシン発現が出芽酵母の代謝に及ぼす影響
    大塚健介, 弘埜陽子, 石井寛子, 石井純, 岡橋伸幸, 戸谷吉博, 原清敬, 松田史生
    2019, 日本生物工学会大会講演要旨集, 71st

  • ロドプシンを利用した大腸菌における光駆動ATP再生の物質生産への応用
    田中涼, 鎌田健太郎, 戸谷吉博, 松田史生, 石井純, 柘植謙爾, 弘埜陽子, 原清敬, 清水浩
    2019, 日本生物工学会大会講演要旨集, 71st

  • 大腸菌による物質生産への光駆動ATP再生の利用に向けて
    田中涼, 鎌田健太郎, 戸谷吉博, 松田史生, 石井純, 柘植謙爾, 弘埜陽子, 原清敬, 清水浩
    2019, 化学工学会年会研究発表講演要旨集(CD-ROM), 84th

  • Combi‐OGAB法を利用した最適コドンの探索
    寺井悟朗, 高橋俊介, 中村朋美, 柘植謙爾, 石井純, 浅井潔
    05 Mar. 2018, 日本農芸化学会大会講演要旨集(Web), 2018, ROMBUNNO.3A29p01 (WEB ONLY), Japanese

  • 光駆動ATP再生系によるVmax細胞の創製
    弘埜陽子, 戸谷吉博, YE Xiaoting, 石井純, 柘植謙爾, 松田史生, 原清敬
    2018, 日本生物工学会大会講演要旨集, 70th

  • 出芽酵母2,3-ブタンジオール生産に向けた不活性型Cas9によるエタノール生合成経路の抑制
    森田啓介, 松田史生, 石井純, 近藤昭彦, 清水浩
    2018, 日本生物工学会大会講演要旨集, 70th

  • ミトコンドリア代謝の抑制による出芽酵母2,3-ブタンジオール生産能力の向上
    森田啓介, 松田史生, 岡本浩二, 石井純, 近藤昭彦, 清水浩
    2018, 日本農芸化学会大会講演要旨集(Web), 2018

  • GPCR拮抗薬のポジティブ検出を可能とする反転型レポーター発現系
    福田 展雄, 石井 純, 本田 真也
    (公社)日本生物工学会, Aug. 2017, 日本生物工学会大会講演要旨集, 平成29年度, 200 - 200, Japanese

  • Kentaro Inokuma, Jun Ishii, Kiyotaka Y. Hara, Masao Mochizuki, Tomohisa Hasunuma, Akihiko Kondo
    AMER SOC MICROBIOLOGY, Feb. 2017, MICROBIOLOGY RESOURCE ANNOUNCEMENTS, 5(5) (5), English
    Others

  • マンナンバイオマスからのエタノール生産:β-マンナナーゼとβ-マンノシダーゼを細胞表層に提示した出芽酵母の開発
    石井純, 岡崎文美, DJOHAN Apridah Cameliawati, 原清敬, 浅井菜々実, ANDRIANI Ade, 寺村浩, KAHAR Prihardi, YOPI, PRASETYA Bambang, 荻野千秋, 近藤昭彦
    2016, 日本農芸化学会大会講演要旨集(Web), 2016

  • 2P-111 Isolation and functional analysis of a novel sesquiterpene synthase gene from Freesia X hybrida 'Ishikawa f1'
    Higuchi,Yuki, Hattan,Jun-ichiro, Ohno,Fumina, Ito,Tomoko, Shibuya,Yurika, Watanabe,Arisa, Ishii,Jun, Kondo,Akihiko, Shindo,Kazutoshi, Misawa,Norihiko
    日本生物工学会, 25 Sep. 2015, 日本生物工学会大会講演要旨集, 67, 202, Japanese

  • 1P-080 Yeast-based biosensor for ligand screening of human nerotensin receptor
    Hashi Hiroki, Nakamura Yasuyuki, Ishii Jun, Kondo Akihiko
    日本生物工学会, 2015, 日本生物工学会大会講演要旨集, 67, 108 - 108, Japanese

  • 2P-110 Isolation and functional analysis of a novel sesquiterpene synthase gene from "Kantsubaki" Camellia hiemalis
    Hattan Jun-ichiro, Shindo Kazutoshi, Ohno Fumina, Higuchi Yuhki, Ishii Jun, Kondo Akihiko, Misawa Norihiko
    日本生物工学会, 2015, 日本生物工学会大会講演要旨集, 67, 202 - 202, Japanese

  • 1P-006 Genome editing system for detecting dimerization of G-protein-coupled receptors
    Hashimoto Takamichi, Nakamura Yasuyuki, Ishii Jun, Kondo Akihiko
    日本生物工学会, 2014, 日本生物工学会大会講演要旨集, 66, 18 - 18, Japanese

  • 3P-226 A display of pH-sensitive fusogenic GALA peptide facilitates endosomal escape for a Bio-nanocapsule
    Nishimura Yuya, Ezawa Ryosuke, Ishii Jun, Ogino Chiaki, Kondo Akihiko
    日本生物工学会, 2014, 日本生物工学会大会講演要旨集, 66, 251 - 251, Japanese

  • 3P-253 Effect of the odorant binding protein on the ligand response of Saccharomyces cerevisiae expressing odorant receptors
    Hori Ayaka, Fukutani Yosuke, Tsukada Satoshi, Sato Ryoichi, Ishii Jun, Kondo Akihiko, Matsunami Hiroaki, Yohda Masafumi
    日本生物工学会, 2014, 日本生物工学会大会講演要旨集, 66, 258 - 258, Japanese

  • 3P-254 Application of the GPCR ligand response detection using split luciferase in budding yeast
    Asakawa Masashi, Fukutani Yosuke, Ishii Jun, Kondo Akihiko, Ozawa Takeaki, Yohda Masafumi
    日本生物工学会, 2014, 日本生物工学会大会講演要旨集, 66, 258 - 258, Japanese

  • 2P-108 Cataloguing terpene synthase genes of fragrant plants : example of Camellia flowers
    Hattan Jun-ichiro, Tagaki Chie, Ohno Fumina, Ishii Jun, Kondo Akihiko, Harimoto Takashi, Misawa Norihiko
    日本生物工学会, 2014, 日本生物工学会大会講演要旨集, 66, 133 - 133, Japanese

  • 2P-220 Cluster analysis of oleaginous yeasts based on their secondary metabolites
    Kojima Motoki, Suzuki Munenori, Harada Hisashi, Misawa Norihiko, Ishii Jun, Kondo Akihiko, Harimoto Takashi
    日本生物工学会, 2014, 日本生物工学会大会講演要旨集, 66, 161 - 161, Japanese

  • Metabolic pathway engineering of yeast Saccharomyces cerevisiae for isobutanol production
    Jun Ishii, Fumio Matsuda, Akihiko Kondo
    WILEY-BLACKWELL, Sep. 2013, YEAST, 30, 210 - 210, English
    Summary international conference

  • A selection system exploiting yeast signal transduction machinery to create desirably affinity-altered protein variants
    Misato Kaishima, Nobuo Fukuda, Jun Ishii, Akihiko Kondo
    WILEY-BLACKWELL, Sep. 2013, YEAST, 30, 182 - 182, English
    Summary international conference

  • 1P-082 High sensitive detection system for ligand screening of human G-protein-coupled receptors by improvement of yeast-based fluorescence reporter assay
    Nakamura Yasuyuki, Ishii Jun, Kondo Akihiko
    日本生物工学会, 2013, 日本生物工学会大会講演要旨集, 65, 38 - 38, Japanese

  • 1P-083 An advanced method to simultaneously analyze the dimerization and signal transduction of G-protein-coupled receptors
    Takemoto Norika, Nakamura Yasuyuki, Ishii Jun, Kondo Akihiko
    日本生物工学会, 2013, 日本生物工学会大会講演要旨集, 65, 38 - 38, Japanese

  • 1P-145 Activation of transhydrogenase-like shunt for isobutanol production in Saccharomyces cerevisiae
    Ishii Jun, Matsuda Fumio, Kondo Takashi, Kondo Akihiko
    日本生物工学会, 2013, 日本生物工学会大会講演要旨集, 65, 54 - 54, Japanese

  • 2P-200 Effect of the accessory proteins on expressing functional odorant receptor with yeast
    Hori Ayaka, Fukutani Yosuke, Ishii Jun, Kondo Akihiko, Matsunami Hiroaki, Yohda Masafumi
    日本生物工学会, 2013, 日本生物工学会大会講演要旨集, 65, 154 - 154, Japanese

  • 3P-174 Development of affibody-displaying bio-nanocapsule for specific delivery to EGFR-expressing cells
    Ezawa Ryosuke, Nishimura Yuya, Ishii Jun, Ogino Chiaki, Kondo Akihiko
    日本生物工学会, 2013, 日本生物工学会大会講演要旨集, 65, 231 - 231, Japanese

  • 神戸大学大学院工学研究科応用化学専攻 生物化学工学研究室
    近藤 昭彦, 荻野 千秋, 田中 勉, 蓮沼 誠久, 石井 純, 原 清敬, 山田 亮祐, 岡井 直子, 工藤 基徳, 佐々木 建吾, 川口 秀夫, 柘植 陽太
    05 Nov. 2012, 化学工学 = Chemical engineering, 76(11) (11), 722 - 722, Japanese

  • 4Ca11 An improved bioluminescence-based signaling assay for odor sensing with a yeast expressing a chimeric olfactory receptor
    Fukutani Yosuke, Ishii Jun, Noguchi Keiichi, Kondo Akihiko, Yohda Masafumi
    日本生物工学会, 2012, 日本生物工学会大会講演要旨集, 64, 194 - 194, Japanese

  • 1Kp05 Design of yeast metabolism for producing next generation biofuels
    Matsuda Fumio, Furusawa Chikara, Ishii Jun, Kondo Takashi, Shimizu Hiroshi, Kondo Akihiko
    日本生物工学会, 2011, 日本生物工学会大会講演要旨集, 63, 68 - 68, Japanese

  • 1Bp13 Production of BIO-1,3-Butanediol from biomass
    Okabayashi Tomohito, Nakajima Takanori, Yamamoto Hiroaki, Kondo Takashi, Ishii Jun, Kondo Akihiko
    日本生物工学会, 2011, 日本生物工学会大会講演要旨集, 63, 27 - 27, Japanese

  • 2Ka04 Reduction of furan derivatives by NADH-dependent Adh1 improves ethanol fermentation using xylose as sole carbon source with Saccharomyces cerevisiae harboring XR-XDH pathway
    Ishii Jun, Yoshimura Kazuya, Hasunuma Tomohisa, Kondo Akihiko
    日本生物工学会, 2011, 日本生物工学会大会講演要旨集, 63, 187 - 187, Japanese

  • 3Ea05 Development of a novel detection system to identify heterodimer forming pairs from human G-protein-coupled receptors
    Nakamura Yasuyuki, Ishii Jun, Kondo Akihiko
    日本生物工学会, 2011, 日本生物工学会大会講演要旨集, 63, 228 - 228, Japanese

  • 3Ea06 A new screening technique for swingably altering affinities of protein
    Kaishima Misato, Fukuda Nobuo, Ishii Jun, Kondo Akihiko
    日本生物工学会, 2011, 日本生物工学会大会講演要旨集, 63, 228 - 228, Japanese

  • 3Ea10 The effect of replacement of N-and C-terminus on the functional expression of mouse olfactory receptor in Saccharomyces cerevisiae
    Fukutani Yosuke, Nakamura Tomoko, Yorozu Maiko, Noguchi Keiichi, Ishii Jun, Kondo Akihiko, Yohda Masafumi
    日本生物工学会, 2011, 日本生物工学会大会講演要旨集, 63, 229 - 229, Japanese

  • Affibody提示Bio-nanocapsuleを用いたHER2発現癌細胞へのタンパク質送達システム
    西村勇哉, 三村和起子, 竹田光一, 石井純, 岡崎文美, 荻野千秋, 近藤昭彦
    2011, 日本生物工学会大会講演要旨集, 63rd

  • 2Ip13 Affibody displaying Bio-nanocapsule for protein therapy to HER2 expressing cancer cells
    Nishimura Yuya, Mimura Wakiko, Takeda Koichi, Ishii Jun, Okazaki Fumiyoshi, Ogino Chiaki, Kondo Akihiko
    日本生物工学会, 2011, 日本生物工学会大会講演要旨集, 63, 180 - 180

  • 2P-1104 A detection system for dimerization of G-protein-coupled receptor using split-ubiquitin and its application for domain analysis
    NAKAMURA Yasuyuki, ISHII Jun, KONDO Akihiko
    日本生物工学会, 2010, 日本生物工学会大会講演要旨集, 22, 34 - 34

  • 2B-Ba05 Single-cell analysis and agonist screening for G-protein-coupled receptors
    Kondo Akihiko, Ishii Jun, Ogino Chiaki
    日本生物工学会, 2010, 日本生物工学会大会講演要旨集, 22, 197 - 197

  • 2P-2051 Efficient fermentation of xylose to ethanol in the presence of furan compound by recombinant Saccharomyces cerevisiae expressing Alcohol dehydrogenase
    YOSHIMURA Kazuya, ISHII Jun, HASUNUMA Tomohisa, KONDO Akihiko
    日本生物工学会, 2010, 日本生物工学会大会講演要旨集, 22, 119 - 119

  • MN-P30 Expression and signaling analyses of human G protein-coupled receptor in yeast(Section X Micro/Nano Technology for Analysis and Cell Manipulation)
    Iguchi Yusuke, Ishii Jun, Tanaka Tsutomu, Kondo Akihiko
    The Society for Biotechnology, Japan, Nov. 2009, Journal of bioscience and bioengineering, 108(1) (1), S164

  • Nobuo Fukuda, Jun Ishii, Tsutomu Tanaka, Akihiko Kondo
    SOC BIOSCIENCE BIOENGINEERING JAPAN, Nov. 2009, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 108, S61 - S62

  • Yusuke Iguchi, Jun Ishii, Tsutomu Tanaka, Akihiko Kondo
    SOC BIOSCIENCE BIOENGINEERING JAPAN, Nov. 2009, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 108, S164 - S164

  • Kazuki Nosaka, Yusuke Miyachi, Jun Ishii, Chiaki Ogino, Akihiko Kondo
    SOC BIOSCIENCE BIOENGINEERING JAPAN, Nov. 2009, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 108, S159 - S160

  • Takanori Tanino, Atsushi Hotta, Tomonori Ito, Jun Ishii, Ryosuke Yamada, Tomohisa Hasunuma, Chiaki Ogino, Naoto Ohmura, Takayuki Ohshima, Akihiko Kondo
    SOC BIOSCIENCE BIOENGINEERING JAPAN, Nov. 2009, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 108, S53 - S53

  • Shota Togawa, Jun Ishii, Tsutomu Tanaka, Akihiko Kondo
    SOC BIOSCIENCE BIOENGINEERING JAPAN, Nov. 2009, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 108, S108 - S108

  • Yasuyuki Nakamura, Jun Ishii, Akihiko Kondo
    SOC BIOSCIENCE BIOENGINEERING JAPAN, Nov. 2009, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 108, S36 - S36

  • 分子ディスプレイによる創薬基盤技術の展開
    芝崎誠司, 石井純, 福田展雄, 近藤昭彦
    2009, 日本薬学会年会要旨集, 129th(1) (1)

  • 酵母ディスプレイ法によるタンパク質の分泌生産/同時回収システム
    芝崎誠司, 川端愛, 石井純, 門之園哲哉, 前田初男, 近藤昭彦, 植田充実
    2008, 日本薬学会年会要旨集, 128th(3) (3)

  • 2Ap01 Biotin-displaying yeast for displaying functional molecule
    MASUNARI Shinsuke, TANAKA Tsutomu, ISHII Jun, FUKUDA Hideki, KONDO Akihiko
    日本生物工学会, 2008, 日本生物工学会大会講演要旨集, 20, 63 - 63

  • タンパク質問相互作用とシグナル伝達解析--酵母細胞を用いたハイスループット技術の開発 (特集 生命化学とバイオ計測)
    石井 純, 田中 勉, 近藤 昭彦
    エヌ・ティー・エス, Jun. 2007, 未来材料, 7(6) (6), 40 - 45

  • 分子ディスプレイ法によるタンパク質の同時生産/回収システムの構築
    芝崎誠司, 川端愛, 石井純, 門之園哲哉, 近藤昭彦, 植田充美
    2007, 生化学

  • 3I09-1 A rapid and high-efficient affinity selection of the surface displayed proteins using temperature-responsive magnetic nanoparticle
    FUKUDA Nobuo, ISHII Jun, FUKUDA Hideki, KONDO Akihiko
    日本生物工学会, 2007, 日本生物工学会大会講演要旨集, 19, 201 - 201

  • Zドメイン表層提示細胞の機能改変と抗体関連分子の精製への応用
    八木俊輔, 石井純, 植田充美, 近藤昭彦, 芝崎誠司
    2006, 日本農芸化学会大会講演要旨集, 2006

  • コンビナトリアル・バイオエンジニアリングによる新しい創薬システム(<特集>コンビナトリアル・バイオエンジニアリング-革新的バイオへの道-)
    石井 純, 近藤 昭彦
    日本生物工学会, 2006, 生物工学会誌 : seibutsu-kogaku kaishi, 84(8) (8), 319 - 321

  • 1J16-2 Development of a novel analysis system for protein interaction by yeast surface display
    FUKUDA Nobuo, ISHII Jun, SHIBASAKI Seiji, FUKUDA Hideki, KONDOU Akihiko
    日本生物工学会, 2006, 日本生物工学会大会講演要旨集, 18, 177 - 177

  • 1H17-2 Development of ligand detection system for heterologous G protein-coupled receptor using yeast
    ISHII Jun, MATSUMURA Shizuka, MORIGUCHI Miyuki, ISHIKURA ATSUSI, IZAWA Keiko, TATEMATSU Kenji, KURODA Syun'ichi, FUKUDA Hideki, KONDO Akihiko
    日本生物工学会, 2006, 日本生物工学会大会講演要旨集, 18, 136 - 136

  • 2A12-3 Development of ligand screening system using the signal transduction in Saccharomyces cerevisiae
    ISHII Jun, MATSUMURA Shizuka, TATEMATSU Kenji, KURODA Syunichi, FUKUDA Hideki, KONDO Akihiko
    日本生物工学会, 2005, 日本生物工学会大会講演要旨集, 17, 62 - 62

  • 2I15-4 Ligand screening system with signal transduction in Saccharomyces cerevisiae
    ISHII Jun, MATSUMURA Shizuka, TATEMATSU Kenji, KURODA Syunichi, FUKUDA Hideki, KONDO Akihiko
    日本生物工学会, 2004, 日本生物工学会大会講演要旨集, 16, 226 - 226

Books And Other Publications

  • ピキア酵母の多重遺伝子欠損による難分泌性抗体タンパク質の生産性向上
    伊藤洋一郎, 石井純, 近藤昭彦
    vol. 81, no. 3, 234-236, バイオサイエンスとインダストリー(バイオインダストリー協会), May 2023

  • 出芽酵母代謝工学株が生産するファイトケミカルの高分解能質量分析による構造確認 –プレニルナリンゲニンの分析–
    岡橋伸幸, 磯貝章太, 石井純, 松田史生
    島津製作所 Application Notes No.72, 島津製作所 Application Notes, Dec. 2021

  • 酵母細胞を用いたG蛋白質共役型受容体リガンド検出システムの開発とメタボライトセンサへの応用
    中村泰之, 近藤昭彦, 石井純
    Precision Medicine(北隆館), Dec. 2021, 第4巻,第13号,60-65

  • 酵母細胞を用いたG蛋白質共役型受容体リガンド検出システムの開発とメタボライトセンサへの応用
    中村泰之, 近藤昭彦, 石井純
    第5巻,第7号,58-63, アグリバイオ(北隆館), Jul. 2021, 第5巻,第7号,58-63

  • 酵母シグナル伝達経路を利用したリガンド検出システムの開発とメタボライトセンサへの応用
    中村泰之, 近藤昭彦, 石井純
    第53巻,第5号,37-40, 月刊細胞(北隆館), May 2021, 第53巻,第5号,37-40

  • 長鎖DNA合成技術による有用物質生産微生物の構築とその課題解決
    柘植謙爾, 石井純, 近藤昭彦
    ゲノム編集技術を応用した製品開発とその実用化 ~研究開発動向・課題解決策・技術予測と市場展望~(第4章 ゲノム編集によるスマートセルインダストリーの技術開発とその課題解決,第7節, 技術情報協会, Feb. 2021

  • 新型コロナで変わる時代の実験自動化・遠隔化「微生物での発酵生産と実験の自動化」
    石井純
    2021年1月号,第39巻,第1号,8-12, 実験医学(羊土社), Jan. 2021, 2021年1月号,第39巻,第1号,8-12

  • 微生物を用いた物質生産とハイスループット微生物構築技術
    石井 純, 西 晶子, 北野 美保, 中村 朋美, 庄司 信一郎, 秀瀨 涼太, 蓮沼 誠久, 近藤 昭彦
    スマートセルインダストリー –微生物細胞を用いた物質生産の展望–(第1編 ハイスループット合成・分析・評価技術,第2章 ハイスループット微生物構築・評価技術,第1節), シーエムシー出版, Jun. 2018

  • G-protein signaling and protein-protein interaction assays for monitoring ligand stimulation and oligomer formation of heterologous GPCRs
    Yasuyuki Nakamura, Akihiko Kondo, Jun Ishii
    Peripheral Membrane Proteins (Chapter 3 "Biosensing techniques in yeast"), InTechOpen, Jun. 2018

  • 生命をつくる –ゲノム構築の時代– 「ゲノム合成の潮流のインパクト 〜微生物による物質生産〜」
    柘植謙爾, 石井純, 荒木通啓, 近藤昭彦
    2018年1月号,第562号,36-41, 現代化学(東京化学同人), Jan. 2018, 2018年1月号,第562号,36-41

  • 低分子抗体医薬の開発展望
    中村泰之, 伊藤洋一郎, 梅津光央, 石井純, 近藤昭彦
    2017年6月号,第34巻,第6号,54-62, 月刊BIO INDUSTRY(シーエムシー出版), Jun. 2017, 2017年6月号,第34巻,第6号,54-62

  • バイオリファイナリーの現状と展望
    蓮沼誠久, 石井純, 荻野千秋, 近藤昭彦
    第53巻,第10号,689-695, 化学と生物(日本農芸化学会誌), Oct. 2015, 第53巻,第10号,689-695

  • 合成生物工学によるモノづくり微生物のデザインに向けて
    石井純, 荒木通啓, 中津井雅彦, 崎濱由梨, 柘植陽太, 蓮沼誠久, 近藤昭彦
    第93巻, 第9号, 523-526, 生物工学会誌, Sep. 2015, 第93巻, 第9号, 523-526

  • ヒドロキシ化合物
    石井 純, 近藤 昭彦
    第Ⅶ編 バイオ科学技術,第26章 バイオマテリアル,第3節, 化学便覧 応用化学編 第7版(丸善出版), Jan. 2014

  • バイオリファイナリー社会に向けた燃料・化学品生産
    石井純, 蓮沼誠久, 松田史生, 近藤昭彦
    第52巻,第4号,249-255, 安全工学(安全工学会 会誌), Aug. 2013, 第52巻,第4号,249-255

  • 革新的なものづくり実現のための「合成生物工学」
    石井純, 蓮沼誠久, 松田史生, 近藤昭彦
    第91巻,第6号,314-318, 生物工学会誌, Jun. 2013, 第91巻,第6号,314-318

  • 酵母による抗体フラグメントおよび抗体様結合性タンパク質の改変技術
    石井 純, 荻野 千秋, 近藤 昭彦
    第Ⅱ編 抗体の改変技術,第4章 親和性の向上,第4節, 次世代医薬開発に向けた抗体工学の最前線(シーエムシー出版), Dec. 2012

  • ノミニケーションノススメ
    石井純
    第89巻,第12号,759, 生物工学会誌, Dec. 2011, 第89巻,第12号,759

  • GPCRと酵母との出会い
    石井純
    内藤財団時報, Sep. 2011, 第88号,47

  • フローサイトメトリーとGFPレポーターによるG蛋白質シグナルのシングルセル解析
    石井 純, 田中 勉, 荻野 千秋, 近藤 昭彦
    第3章 細胞内生体分子群の実測定量解析,第4節, シングルセル解析の最前線 (シーエムシー出版), Mar. 2010

  • タンパク質間相互作用とシグナル伝達解析¬ −酵母細胞を用いたハイスループット技術の開発-−
    石井純, 田中勉, 近藤昭彦
    2007年6月号,40-45, 未来材料(出版社NTS), Jun. 2007, 2007年6月号,40-45

  • 酵母細胞内シグナル定量解析の創薬への応用
    石井 純, 近藤 昭彦
    (第3章 細胞内生体分子群の動態シグナルの解析,第6節), 細胞定量解析の最前線 −ライフサーベイヤ構築に向けて−(シーエムシー出版), Dec. 2006

  • コンビナトリアル・バイオエンジニアリングによる新しい創薬システム
    石井純, 近藤昭彦
    第84巻,第8号,319-321, 生物工学会誌, Aug. 2006, 第84巻,第8号,319-321

Lectures, oral presentations, etc.

  • 大腸菌によるレチナールの生産
    雨宮彩恵, 弘埜陽子, 松田史生, 戸谷吉博, 石井純, 原清敬
    日本農芸化学会2024年度大会, Mar. 2024, 日本農芸化学会, 東京農工大学  世田谷キャンパス

  • T7ファージ由来の転写系を利用した酵母遺伝子スイッチの開発
    堀智彦, 冨永将大, 梶亘佑, 近藤昭彦, 石井純
    神戸大学 研究基盤センター 若手フロンティア研究会2023, Dec. 2023, 神戸大学 研究基盤センター, 神戸大学 百年記念館

  • Development of genetic engineering tools for synthetic biology and cell factories in yeasts
    Jun Ishii
    2023 Asian Synthetic Biology Association (ASBA) Meeting (ASBA2023), Dec. 2023, Asian Synthetic Biology Association (ASBA), Awaji Yumebutai International Conference Center, Japan

  • Development of antibody binding protein-immobilized functional microhydrogels with cell sorting capability
    Noe Inomoto, Masahiro Tominaga, Yoichiro Ito, Jun Ishii, Akihiko Kondo, Noriho Kamiya
    The 34th International Symposium on Chemical Engineering (ISChE2023), Dec. 2023, The Society of Chemical Engineers, Japan (SCEJ) & The Korean Institute of Chemical Engineers (KIChE), Gyeongju, South Korea

  • バクテリオファージ由来の転写系を利用した酵母遺伝子スイッチの構築
    堀智彦, 冨永将大, 梶亘佑, 近藤昭彦, 石井純
    日本農芸化学会関西支部第528回講演会, Dec. 2023, 日本農芸化学会関西支部, 神戸大学 六甲台キャンパス

  • データ駆動型の次世代微生物進化育種
    石井純
    2023年度CREST「データ駆動・AI駆動を中心としたデジタルトランスフォーメーションによる生命科学研究の革新」領域キックオフシンポジウム, Nov. 2023, 科学技術振興機構(JST), AP市ヶ谷

  • バクテリオファージ由来転写系を利用した酵母遺伝子スイッチの開発
    堀智彦, 冨永将大, 梶亘佑, 近藤昭彦, 石井純
    第3回神戸大学先端バイオ工学研究センター成果発表会, Sep. 2023, 神戸大学先端バイオ工学研究センター, 神戸大学 百年記念館

  • 高発現性の酵母人工誘導プロモータの開発
    冨永将大, 伊藤洋一郎, 近藤昭彦, 石井純
    第3回神戸大学先端バイオ工学研究センター成果発表会, Sep. 2023, 神戸大学先端バイオ工学研究センター, 神戸大学 百年記念館六甲ホール

  • TECAN Fluentを用いた分泌生産タンパク質の多種同時自動精製
    片山健太, 中村泰之, 伊藤洋一郎, 石井純, 近藤昭彦
    Laboratory Automation Developers Conference 2023 (LADEC2023), Sep. 2023, ラボラトリーオートメーション研究会, 理化学研究所 生命機能科学研究センター

  • 光駆動プロトンポンプを利用した大腸菌の酸耐性の強化と有用物質生産への応用
    小林祐摩, 佐野海瑚人, 弘埜陽子, 松田史生, 石井純, 原清敬, 清水浩, 戸谷吉博
    第75回日本生物工学会大会, Sep. 2023, 日本生物工学会, 名古屋大学東山キャンパス

  • CHO細胞における二重特異性scFv抗体の発現と分子特性評価
    福間奈々子, 沖田大和, 天羽宏枝, 伊藤洋一郎, 石井純, 近藤昭彦, 梅津光央, 鬼塚正義
    第75回日本生物工学会大会, Sep. 2023, 日本生物工学会, 名古屋大学東山キャンパス

  • 細胞選別能を有する機能性微小ハイドロゲルの創製
    井元乃絵, 冨永将大, 伊藤洋一郎, 石井純, 近藤昭彦, 神谷典穂
    第75回日本生物工学会大会, Sep. 2023, 日本生物工学会, 名古屋大学東山キャンパス

  • バクテリオファージ由来RNAポリメラーゼを利用した酵母遺伝子スイッチの開発
    堀智彦, 冨永将大, 梶亘佑, 近藤昭彦, 石井純
    第75回日本生物工学会大会, Sep. 2023, 日本生物工学会, 名古屋大学東山キャンパス

  • 酵母の高発現性人工誘導プロモータの開発
    冨永将大, 伊藤洋一郎, 近藤昭彦, 石井純
    第75回日本生物工学会大会, Sep. 2023, 日本生物工学会, 名古屋大学東山キャンパス

  • ピキア酵母におけるターミネーター置換での遺伝子発現量制御
    伊藤洋一郎, 寺井悟朗, 石上美佐, 橋場倫子, 中村泰之, 番場崇弘, 雲北涼太, 蓮沼誠久, 浅井潔, 石井純, 近藤昭彦
    第75回日本生物工学会大会, Sep. 2023, 日本生物工学会, 名古屋大学東山キャンパス

  • 微生物代謝の理解と最適化に向けたバイオDX基盤の構築
    石井純
    第75回日本生物工学会大会 シンポジウム(生命科学研究の革新に向けたバイオDXの挑戦), Sep. 2023, 日本生物工学会/JST・CREST, 名古屋大学東山キャンパス

  • Light-driven ATP regeneration for enhancing 3-hydroxypropionic acid and isoprenol productions in Escherichia coli
    Yoshihiro Toya, Mikoto Sano, Ryo Tanaka, Kentaro Kamata, Fumio Matsuda, Jun Ishii, Yoko Hirono-Hara, Kiyotaka Y. Hara, Hiroshi Shimizu
    Metabolic Engineering 15, American Institute of Chemical Engineers (AIChE), Marina Bay Sands, Singapore

  • 酵母における合成生物学基盤の開発と物質生産への展開
    石井純
    日本農芸化学会2023年度大会 シンポジウム(微生物によるものづくり研究の最前線 ~宿主機能開発から社会実装に向けた取り組みまで), Mar. 2023, 日本農芸化学会, オンライン

  • メチロトローフ酵母を用いた異なる炭素源からの乳酸生産
    鈴木創太, 廣畑凌治, 中村泰之, 伊藤洋一郎, 北野美穂, 冨永将大, 中曽根光, 近藤昭彦, 石井純
    第25回化学工学会学生発表会, Mar. 2023, 化学工学会, オンライン

  • 光エネルギーで有用物質を高生産する微生物の開発
    戸谷吉博, 弘埜(原)陽子, 平山英伸, 鎌田健太郎, 田中涼, 佐野海瑚人, 北村さや香, 大塚健介, 吉住玲, 角田聡, 菊川寛史, 神取秀樹, 清水浩, 松田史生, 石井純, 原清敬
    日本生体エネルギー研究会 第48回討論会, Dec. 2022, 日本生体エネルギー研究会, 京都大学北部キャンパス 益川ホール

  • バクテリア転写因子CamRを用いたボルネオール生産酵母のスクリーニング系の構築
    川上和真, 冨永将大, 小川ひろ, 能崎健太, 近藤昭彦, 石井純
    日本農芸化学会関西支部第523回講演会, Dec. 2022, 日本農芸化学会関西支部, オンライン

  • 動物細胞を利用したタンデム型二重特異性scFv抗体の製造適合性評価
    福間奈々子, 天羽宏枝, 伊藤洋一郎, 石井純, 近藤昭彦, 梅津光央, 鬼塚正義
    第1回抗体学会学術大会, Nov. 2022, 日本抗体学会, 鹿児島大学学習交流プラザ・オンライン併催

  • データ駆動型の次世代微生物進化育種
    石井純
    2022年度CREST「バイオDX」領域キックオフシンポジウム, Nov. 2022, 科学技術振興機構(JST), JST東京本部別館・オンライン併催

  • 大腸菌におけるロドプシンの光駆動プロトンポンプが弱酸耐性に及ぼす影響
    小林祐摩, 佐野海瑚人, 弘埜陽子, 松田史生, 石井純, 原清敬, 戸谷吉博, 清水浩
    第74回日本生物工学会大会(100周年記念大会), Oct. 2022, 日本生物工学会, オンライン

  • 微生物ものづくりのための合成生物学と実験自動化:バイオエコノミーの実現に向けて
    石井純
    第74回日本生物工学会大会(100周年記念大会)ランチタイムセミナー, Oct. 2022, 日本生物工学会, オンライン

  • ATP浪費による出芽酵母バイオアルコール生産能の向上
    谷田部楓太, 岡橋伸幸, 清家泰介, 石井純, 松田史生
    第74回日本生物工学会大会(100周年記念大会), Oct. 2022, 日本生物工学会, オンライン

  • 有用タンパク質の高生産化に向けた新たなピキア酵母株の開発方針
    伊藤洋一郎, 石上美佐, 寺井悟朗, 中村泰之, 橋場倫子, 西輝之, 中澤光, 蓮沼誠久, 浅井潔, 梅津光央, 石井純, 近藤昭彦
    第74回日本生物工学会大会(100周年記念大会), Oct. 2022, 日本生物工学会, オンライン

  • 酵母におけるスクアレン生産の呈色評価系の構築
    三ツ橋拓海, 冨永将大, 宮崎敬太, 近藤昭彦, 石井純
    第2回神戸大学先端バイオ工学研究センター成果発表会, Oct. 2022, 神戸大学先端バイオ工学研究センター, 神戸大学 百年記念館

  • ピキア酵母を用いた高機能な低分子抗体のスクリーニング系の確立
    中曽根光, 伊藤洋一郎, 近藤昭彦, 石井純
    第2回神戸大学先端バイオ工学研究センター成果発表会, Oct. 2022, 神戸大学先端バイオ工学研究センター, 神戸大学 百年記念館

  • 植物由来テルペン合成酵素を用いたボルネオール生産酵母の開発
    川上和真, 冨永将大, 小川ひろ, 能崎健太, 近藤昭彦, 石井純
    第2回神戸大学先端バイオ工学研究センター成果発表会, Oct. 2022, 神戸大学先端バイオ工学研究センター, 神戸大学 百年記念館

  • メタボローム分析によるバイオアルコール生産能改変:出芽酵母の代謝律速点の推定
    杉村政彦, 清家泰介, 岡橋伸幸, 石井純, 松田史生
    第16回メタボロームシンポジウム, Sep. 2022, 第16回メタボロームシンポジウム実行委員会, 鶴岡メタボロームキャンパス

  • 代謝工学と合成生物学によるモノづくり微生物の開発
    石井純
    2021年度膜工学春季講演会・膜工学サロン, Mar. 2022, 神戸大学先端膜工学研究センター/一般社団法人先端膜工学研究機構, オンライン

  • バイオエコノミー社会実現に向けた合成生物学の基盤開発と微生物での物質生産への展開
    石井純
    SONY “Life Science Spring Webinar 2022”, Mar. 2022, SONY, オンライン

  • ロドプシンを利用した光駆動ATP再生によるイソプレノールおよび3ヒドロキシプロピオン酸生産の向上
    戸谷吉博, 佐野海瑚人, 田中涼, 鎌田健太郎, 松田史生, 石井純, 弘埜陽子, 原清敬, 清水浩
    化学工学会第87年会, Mar. 2022, 化学工学会, 日本

  • 酵母シグナル伝達機構を利用したタンパク質間相互作用分子探索および改変体創出
    伊藤沙也加, 浅間梨々花, 杉本美帆, 桂田康平, 近藤昭彦, 石井純
    化学工学会第87年会, Mar. 2022, 化学工学会, オンライン

  • ロドプシンによる光駆動プロトンポンプが大腸菌の弱酸耐性に及ぼす影響
    小林祐摩, 佐野海瑚人, 弘埜陽子, 松田史生, 石井純, 原清敬, 戸谷吉博, 清水浩
    化学工学会第87年会, Mar. 2022, 化学工学会, オンライン

  • 微生物生産能向上を目的とした微生物ロドプシンの改変
    佐藤良樹, 川崎真由, 弘埜陽子, 菊川寛史, 中野祥吾, 石井純, 松田史生, 戸谷吉博, 伊藤創平, 原清敬
    日本農芸化学会2022年度大会, Mar. 2022, 日本農芸化学会, オンライン

  • 酵母シグナル伝達機構を利用したタンパク質間相互作用分子探索および改変体創出
    伊藤沙也加, 浅間梨々花, 杉本美帆, 桂田康平, 近藤昭彦, 石井純
    日本農芸化学会関西支部第519回講演会, Feb. 2022, 日本農芸化学会関西支部, オンライン

  • 微生物モノづくりにおける合成生物学と実験自動化:バイオエコノミー社会の実現に向けて
    石井純
    第12回スクリーニング学研究会(企業セミナー:テカンジャパン), Nov. 2021, スクリーニング学研究会, オンライン

  • データ駆動型の次世代微生物進化育種に向けて
    石井純
    バイオDXの最前線 CREST 「データ駆動型・AI駆動を中心としたデジタルトランスフォーメーションによる生命科学研究の革新」キックオフシンポジウム, Nov. 2021, 科学技術振興機構(JST)/理化学研究所 生命機能科学研究センター, オンラインn

  • いっぱいデータを出したいがため微生物の組換え実験を自動化する
    石井純
    Laboratory Automation月例勉強会 / 2021.10,, Oct. 2021, ラボラトリーオートメーション研究会, オンライン

  • 光駆動プロトンポンプを発現させたメバロン酸生産大腸菌の代謝解析
    松山千夏, 清家泰介, 岡橋伸幸, 戸谷吉博, 弘埜陽子, 平山英伸, 石井純, 清水浩, 原清敬, 松田史生
    第73回日本生物工学会大会, Oct. 2021, 日本生物工学会, オンライン

  • H+ポンプ能向上を目的とした微生物ロドプシンの改変
    佐藤良樹, 川崎真由, 弘埜陽子, 菊川寛史, 中野祥吾, 石井純, 松田史生, 戸谷吉博, 伊藤創平, 原清敬
    第73回日本生物工学会大会, Oct. 2021, 日本生物工学会, オンライン

  • 光エネルギーを利用した大腸菌におけるメバロン酸のイソプレノールへの変換
    佐野海瑚人, 田中涼, 鎌田健太郎, 松田史生, 石井純, 弘埜陽子, 原清敬, 戸谷吉博, 清水浩
    第73回日本生物工学会大会, Oct. 2021, 日本生物工学会, オンライン

  • Xanthophyllomyces dendrorhousによるレチナールの生産
    藤野紗代, 景山裕也, 弘埜陽子, 菊川寛史, 松田史生, 戸谷吉博, 石井純, 原清敬
    第73回日本生物工学会大会, Oct. 2021, 日本生物工学会, オンライン

  • 進化工学による酵母遺伝子スイッチの開発とテルペノイドセンサへの応用
    冨永将大, 川上和真, 小川ひろ, 能崎健太, 近藤昭彦, 石井純
    第1回神戸大学先端バイオ工学センター成果発表会, Oct. 2021, 先端バイオ工学センター, オンライン

  • ピキア酵母による低分子抗体高生産化
    中村泰之, 佐々木大介, 伊藤洋一郎, 近藤昭彦, 石井純
    第1回神戸大学先端バイオ工学センター成果発表会, Oct. 2021, 先端バイオ工学センター, オンライン

  • 分泌タンパク質精製のハイスループット自動化
    堀川拓真, 伊藤洋一郎, 近藤昭彦, 石井純
    第1回神戸大学先端バイオ工学センター成果発表会, Oct. 2021, 先端バイオ工学センター, オンライン

  • メタノール資化性酵母Pichia pastorisによる有用化合物生産プロセス開発
    廣畑凌治, 中村泰之, 近藤昭彦, 石井純
    第1回神戸大学先端バイオ工学センター成果発表会, Oct. 2021, 神戸大学先端バイオ工学センター, オンライン

  • メタノール資化性酵母Pichia pastorisにおけるゲノム編集技術の開発
    梶川耕介, 伊藤洋一郎, 山田健人, 南部由美子, 西田敬二, 近藤昭彦, 石井純
    第1回神戸大学先端バイオ工学センター成果発表会, Oct. 2021, 神戸大学先端バイオ工学センター, オンライン

  • 大腸菌における有用物質生産に適した鉄硫黄タンパク質発現系の構築
    白河原拓実, 秀瀬涼太, 酒井香奈江, 冨永将大, 近藤昭彦, 石井純
    第1回神戸大学先端バイオ工学センター成果発表会, Oct. 2021, 神戸大学先端バイオ工学センター, オンライン

  • Development of engineering tools for methylotrophic yeast Pichia pastoris and their applications to small antibody secretory production
    Akihiko Kondo, Yoichiro Ito, Yasuyuki Nakamura, Daisuke Sasaki, Jun Ishii
    The 15th International Congress on Yeasts (ICY15), online

  • 酵母における合成生物学のための基盤ツール開発と有用物質生産細胞育種への応用
    石井純
    酵母研究会第89回講演会, Aug. 2021, 酵母研究会, オンライン

  • 酵母を用いたセルファクトリーとスマートセル創出に向けた基盤技術開発
    石井純
    日本化学会第101年会春季年会 イノベーション共創プログラム(CIP)(スマートセルインダストリーという未来), Mar. 2021, 日本化学会, オンライン

  • 酵母におけるスクアレン生合成経路の改変および下流モノオキシゲナーゼの発現調節
    宮崎敬太, 三井靖雅, 旗谷章子, 冨永将大, 近藤昭彦, 石井純
    日本農芸化学会関西支部第513回講演会, Nov. 2020, 日本農芸化学会関西支部, オンライン

  • ドーパミン発酵生産性を簡易的に評価するGPCRメタボライトセンサの開発
    浅間梨々花, 中村泰之, 中村朋美, 近藤昭彦, 石井純
    日本農芸化学会関西支部第513回講演会, Nov. 2020, 日本農芸化学会関西支部, オンライン

  • Development of engineering tools for methylotrophic yeast Pichia pastoris and their applications to small antibody secretory production
    Akihiko Kondo, Yoichiro Ito, Yasuyuki Nakamura, Yuya Nishimura, Daisuke Sasaki, Teruyuki Nishi, Goro Terai, Kiyoshi Asai, Jun Ishii
    International Union of Microbiological Societies 2020 Congresses (IUMS 2020), online

  • 自動分注装置を用いた微生物形質転換システムの構築
    石井純, 舘野雄紀
    Laboratory Automation Developers Conference 2020(LADEC2020), Oct. 2020, ラボラトリーオートメーション研究会, オンライン

  • Development of dopamine sensor using G protein-coupled receptor and its application to screening of dopamine producers
    Ririka Asama, Yasuyuki Nakamura, Tomomi Nakamura, Akihiko Kondo, Jun Ishii
    化学工学会第51回秋季大会, Sep. 2020, 化学工学会, オンライン

  • 食品中の有毒物質を検出するためのパトロール酵母の開発
    蘇九龍, 中村泰之, 北口哲也, 石井純, 近藤昭彦, 小林典裕, 三宅司郎, 上田宏
    第93回日本生化学会大会, Sep. 2020

  • 低毒性とゲル化能を両立した短鎖ペプチドゲル化剤
    Kartika Restu Witta, 山本翔太, 石井純, 青井貴之, 丸山達生
    化学工学会第85年会, Mar. 2020, 化学工学会

  • Analysis of deltarhodopsin mutant
    佐藤良樹, 弘埜陽子, 石井純, 柘植謙爾, 松田史生, 戸谷吉博, 原清敬
    日本農芸化学会2020年度大会, Mar. 2020, 日本農芸化学会

  • Improvement of bioproduction with ATP recycling by delta-rhodopsin
    塩澤優希, 弘埜陽子, 石井純, 柘植謙爾, 松田史生, 戸谷吉博, 原清敬
    日本農芸化学会2020年度大会, Mar. 2020, 日本農芸化学会

  • 進化デザインによる新規テルペノイドセンサの開発
    冨永将大, 小川ひろ, 能崎健太, 近藤昭彦, 石井純
    日本農芸化学会関西支部 支部例会(第511回講演会), Dec. 2019

  • メラトニン生産性を簡便かつハイスループットに評価するメタボライトセンサの開発
    浅間梨々花, 田畑琢也, 中村泰之, 中村朋美, 加藤寛子, 近藤昭彦, 石井純
    日本農芸化学会関西支部 支部例会(第511回講演会), Dec. 2019

  • 新規な3機能性融合マーカーを用いた酵母遺伝子スイッチの組織的開発
    能崎健太, 冨永将大, 梅野太輔, 近藤昭彦, 石井純
    日本農芸化学会関西支部 支部例会(第511回講演会), Dec. 2019

  • Construction of metabolite sensor using yeast signal transduction machinery for monitoring melatonin production
    Jun Ishii, Takuya Tabata, Tomomi Nakamura, Hiroko Kato, Ririka Asama, Yasuyuki Nakamura, Akihiko Kondo
    The 10th Symposium on ​Innovative Bioproduction Taichung (iBioT2019), Nov. 2019, Tunghai University, Taichung

  • Construction of melatonin metabolite sensor using yeast signal transduction machinery
    Jun Ishii, Takuya Tabata, Tomomi Nakamura, Hiroko Kato, Ririka Asama, Yasuyuki Nakamura, Akihiko Kondo
    2019 Asian Synthetic Biology Association (ASBA) Meeting (ASBA2019), Oct. 2019, JW Marriott Hotel Chengdu, Chengdu

  • 合成生物学とバイオエコノミー
    石井純
    第5回Laboratory Automation勉強会, Oct. 2019, ラボラトリーオートメーション研究会, 神戸大学 百年記念館 六甲ホール

  • Feasibility study of the method to obtain peptide agonists for G protein-coupled receptors using water-in-oil microdroplets
    Anna Matsueda, Takashi Sakurai, Ryo Iizuka, Yasuyuki Nakamura, Jun Ishii, Akihiko Kondo, Dong Hyun Yoon, Tetsushi Sekiguchi, Shuichi Shoji, Soichiro Tsuda, Takashi Funatsu
    第57回日本生物物理学会年会, Sep. 2019, 日本生物物理学会, 宮崎県・シーガイアコンベンションセンター

  • 大腸菌・酵母の半自動形質転換システムとハイスループット生産性評価
    石井 純
    第92回日本生化学会大会(シンポジウム「実験自動化の今」), Sep. 2019, 日本生化学会, パシフィコ横浜

  • 陽性/陰性選択マーカーと蛍光タンパク質から成る3機能性新規融合タンパク質を用いた酵母遺伝子スイッチの開発
    能崎 健太, TOMINAGA MASAHIRO, 梅野 大輔, KONDO AKIHIKO, ISHII JUN
    第71回日本生物工学会大会, Sep. 2019, 日本生物工学会, 岡山大学 津島キャンパス

  • デルタロドプシン発現が出芽酵母の代謝に及ぼす影響
    大塚 健介, 原 清敬, 石井 寛子, ISHII JUN, 岡橋 伸幸, 戸谷 吉博, 松田 史生
    第71回日本生物工学会大会, Sep. 2019, 日本生物工学会, 岡山大学 津島キャンパス

  • Pichia pastorisにおいて分泌シグナル配列の1アミノ酸置換はタンパク質分泌生産量を劇的に増大させる
    ITO YOICHIRO, 石上 美佐, 橋場 倫子, NAKAMURA YASUYUKI, HASUNUMA TOMOHISA, ISHII JUN, KONDO AKIHIKO
    第71回日本生物工学会大会, Sep. 2019, 日本生物工学会, 岡山大学 津島キャンパス

  • ロドプシンを利用した大腸菌における光駆動ATP再生の物質生産への応用
    田中 涼, 鎌田 健太郎, 戸谷 吉博, 松田 史生, ISHII JUN, TSUGE KENJI, 弘埜 陽子, 原 清敬, 清水 浩
    第71回日本生物工学会大会, Sep. 2019, 日本生物工学会, 岡山大学 津島キャンパス

  • 光駆動ATP再生系による出芽酵母型Vmax細胞の創製
    弘埜 陽子, 塩澤 優希, 戸谷 吉博, ISHII JUN, TSUGE KENJI, 松田 史生, 菊川 寛史, 原 清敬
    酵母遺伝学フォーラム第52回研究報告会, Sep. 2019, 酵母遺伝学フォーラム, 静岡市清水文化会館マリナート

  • 分泌タンパク質精製のハイスループット自動化
    HORIKAWA TAKUMA, ITO YOICHIRO, ISHII JUN, KONDO AKIHIKO
    生物工学若手研究者の集い 夏のセミナー2019, Jul. 2019, 日本生物工学会 生物工学若手研究者の集い(若手会), 琵琶湖国定公園 近江白浜 政府登録旅館 白浜荘

  • 酵母を用いた癌標的膜タンパク質に対する相互作用阻害分子のスクリーニング系構築
    KATSURADA KOUHEI, SUGIMOTO MIHO, KAISHIMA MISATO, TOMINAGA MASAHIRO, KONDO AKIHIKO, ISHII JUN
    生物工学若手研究者の集い 夏のセミナー2019, Jul. 2019, 日本生物工学会 生物工学若手研究者の集い(若手会), 琵琶湖国定公園 近江白浜 政府登録旅館 白浜荘

  • 抗体生産性に関わる遺伝子探索のためのPichia pastorisゲノム編集技術の構築
    YAMADA KENTO, NANBU-NISHIDA YUMIKO, ITO YOICHIRO, NISHIDA KEIJI, ISHII JUN, KONDO AKIHIKO
    生物工学若手研究者の集い 夏のセミナー2019, Jul. 2019, 日本生物工学会 生物工学若手研究者の集い(若手会), 琵琶湖国定公園 近江白浜 政府登録旅館 白浜荘

  • メラトニン発酵生産性を簡便に評価する酵母メタボライトセンサの開発
    ASAMA RIRIKA, TABATA TAKUYA, NAKAMURA YASUYUKI, KATO HIROKO, KONDO AKIHIKO, ISHII JUN
    生物工学若手研究者の集い 夏のセミナー2019, Jul. 2019, 日本生物工学会 生物工学若手研究者の集い(若手会), 琵琶湖国定公園 近江白浜 政府登録旅館 白浜荘

  • ノカルジチオシン二次代謝生合成遺伝子クラスターの大腸菌発現系構築
    MATSUMOTO KAZUKI, SAKAI KANAE, TSUGE KENJI, ISHII JUN, KONDO AKIHIKO
    生物工学若手研究者の集い 夏のセミナー2019, Jul. 2019, 日本生物工学会 生物工学若手研究者の集い(若手会), 琵琶湖国定公園 近江白浜 政府登録旅館 白浜荘

  • デルタロドプシンを用いた光エネルギーによるATP再生能力強化酵母株の構築
    大塚 健介, 弘埜 陽子, 原 清敬, 石井 寛子, ISHII JUN, 岡橋 伸幸, 戸谷 吉博, 松田 史生
    生物工学若手研究者の集い 夏のセミナー2019, Jul. 2019, 日本生物工学会 生物工学若手研究者の集い(若手会), 琵琶湖国定公園 近江白浜 政府登録旅館 白浜荘

  • 光駆動ATP再生系によるVmax細胞の創製
    弘埜 陽子, 戸谷 吉博, ISHII JUN, TSUGE KENJI, 松田 史生, 原 清敬
    日本農芸化学会2019年度大会, Mar. 2019, 日本農芸化学会, 東京農業大学 世田谷キャンパス

  • 大腸菌による物質生産への光駆動ATP再生の利用に向けて
    田中 涼, 鎌田 健太郎, 戸谷 吉博, 松田 史生, ISHII JUN, TSUGE KENJI, 弘埜 陽子, 原 清敬, 清水 浩
    化学工学会第84年会, Mar. 2019, 化学工学会, 芝浦工業大学 豊洲キャンパス

  • Gタンパク質共役型受容体ペプチドアゴニスト創出法のハイスループット化
    松枝 杏奈, 櫻井 貴志, 飯塚 怜, NAKAMURA YASUYUKI, ISHII JUN, KONDO AKIHIKO, 井口 彩香, 尹 棟鉉, 関口 哲志, 庄子 習一, 藤村 祐, 赤木 仁, 石毛 真行, 船津 高志
    日本薬学会第139年会, Mar. 2019, 日本薬学会, 幕張メッセ

  • DNAシャッフリングを利用した出芽酵母におけるコドン最適化ルールの抽出
    寺井 悟朗, 高橋 俊介, 中村 朋美, 柘植 謙爾, 石井 純, 浅井 潔
    第41回日本分子生物学会年会, Nov. 2018, 日本分子生物学会, パシフィコ横浜

  • Yeast-based in vivo metabolite sensor using signal transduction machinery
    Jun Ishii, Takuya Tabata, Yasuyuki Nakamura, Akihiko Kondo
    2018 Asian Synthetic Biology Association (ASBA) Meeting (ASBA2018), Nov. 2018, Hyatt Regency Jeju

  • Construction of a stable, autonomously replicating plasmid vector containing Pichia pastoris centromeric DNA
    NAKAMURA YASUYUKI, Teruyuki Nishi, Risa Noguchi, ITO YOICHIRO, Toru Watanabe, Tozo Nishiyama, Shimpei Aikawa, Tomohisa Hasunuma, ISHII JUN, Yoshihiko Yasohara, Akihiko Kondo
    The Symposium on Biorefinery and Biprocess Topics, 2018 (iBio-N 2018), Nov. 2018, Nanjing, China

  • 物質生産宿主としての酵母の代謝経路改変とゲノム改変に向けた合成生物工学的手法開発
    Jun Ishii
    第70回日本生物工学会大会(シンポジウム「新時代の物質生産宿主開発の方法論: ゲノムを大規模に編集する。代謝計測から設計図を書く。」), Sep. 2018, 日本生物工学会, 関西大学 千里山キャンパス

  • 出芽酵母2,3-ブタンジオール生産に向けた不活性型Cas9によるエタノール生合成経路の抑制
    森田 啓介, 松田 史生, 石井 純, 近藤 昭彦, 清水 浩
    第70回日本生物工学会大会, Sep. 2018, 関西大学 千里山キャンパス

  • 酵母細胞における生理活性物質メラトニンをモニタリングするためのGタンパク質共役型受容体(GPCR)を用いたメタボライトセンサの開発
    田畑 琢也, NAKAMURA YASUYUKI, ISHII JUN, 加藤 寛子, KONDO AKIHIKO
    第70回日本生物工学会大会, Sep. 2018, 日本生物工学会, 関西大学 千里山キャンパス

  • 光駆動ATP再生系によるVmax細胞の創製
    弘埜 陽子, 戸谷 吉博, 叶 暁亭, ISHII JUN, TSUGE KENJI, 松田 史生, 原 清敬
    第70回日本生物工学会大会, Sep. 2018, 日本生物工学会, 関西大学 千里山キャンパス

  • 遺伝子組換え効率向上に向けたDNAリガーゼIV欠損Pichia pastoris株の開発
    ITO YOICHIRO, 渡邉 徹, 藍川 晋平, 西 輝之, 西山 陶三, NAKAMURA YASUYUKI, HASUNUMA TOMOHISA, 八十原 良彦, ISHII JUN, KONDO AKIHIKO
    第70回日本生物工学会大会, Sep. 2018, 日本生物工学会, 関西大学 千里山キャンパス

  • Pichia pastorisにおける自律複製型ベクターを用いた効率的なDNAマルチアセンブル法
    西 輝之, 西山 陶三, 山路 大樹, 玉井 雅也, 渡邉 徹, NAKAMURA YASUYUKI, ITO YOICHIRO, ISHII JUN, KONDO AKIHIKO, 八十原 良彦
    第70回日本生物工学会大会, Sep. 2018, 日本生物工学会, 関西大学 千里山キャンパス

  • Pichia pastorisにおけるセントロメアDNA配列を用いた自律複製型プラスミドベクターの開発
    NAKAMURA YASUYUKI, 西 輝之, 野口 理紗, ITO YOICHIRO, 渡邉 徹, 西山 陶三, 藍川 晋平, HASUNUMA TOMOHISA, ISHII JUN, 八十原 良彦, KONDO AKIHIKO
    第70回日本生物工学会大会, Sep. 2018, 日本生物工学会, 関西大学 千里山キャンパス

  • ハイスループット微生物構築・評価技術の開発
    ISHII JUN, 西 晶子, 北野 美保, 中村 朋美, KATO HIROKO, SHOUJI SHINICHIROU, HIDESE RYOTA, 梅野 太輔
    第20回 新産業技術促進検討会, Jul. 2018, モノづくり日本会議/日刊工業新聞社

  • 発酵制御に向けたグルコースを感知するin vivoバイオセンサーの構築
    橋 弘樹, Yasuyuki Nakamura, Akihiko Kondo, Jun Ishii
    第3回デザイン生命工学研究会, Mar. 2018, デザイン生命工学研究, 今帰仁村コミュニティセンター

  • 出芽酵母における代謝経路デザインと高級アルコール生産
    Jun Ishii, 森田 啓介, 伊藤 賢吾, 加藤 寛子, 木下 翔平, 旗谷 章子, 清水 浩, Akihiko Kondo, 松田 史生
    第3回デザイン生命工学研究会, Mar. 2018, デザイン生命工学研究, 今帰仁村コミュニティセンター

  • 麹菌を用いた遊離型高度不飽和脂肪酸の生産化
    玉野 孝一, Robert Sidney Cox III, Kenji Tsuge, 三浦 愛, 伊藤 あやの, Jun Ishii, 田村 具博, Akihiko Kondo, 町田 雅之
    日本農芸化学会2018年度大会, Mar. 2018, 日本農芸化学会, 名城大学 天白キャンパス

  • 酵母シグナル伝達機構を利用した膜タンパク質に対する相互作用を阻害する分子スクリーニングシステムの構築
    杉本 美帆, 松原 大希, 海嶋 美里, 福田 展雄, Akihiko Kondo, Jun Ishii
    第3回デザイン生命工学研究会, Mar. 2018, デザイン生命工学研究, 今帰仁村コミュニティセンター

  • ミトコンドリア代謝の抑制による出芽酵母2,3-ブタンジオール生産能力の向上
    森田 啓介, 松田 史生, 岡本 浩二, Jun Ishii, Akihiko, 清水 浩
    日本農芸化学会2018年度大会, Mar. 2018, 日本農芸化学会, 名城大学 天白キャンパス

  • P. pastorisにおける自律複製型ベクターを用いた効率的なDNAマルチアセンブル法
    西 輝之, 山路 大樹, 玉井 雅也, 渡邉 徹, 西山 陶三, NAKAMURA YASUYUKI, ITO YOICHIRO, ISHII JUN, KONDO AKIHIKO, 八十原 良彦
    第12回日本ゲノム微生物学会年会, Mar. 2018, 日本ゲノム微生物学会, 京都大学 桂キャンパス

  • Combi-OGAB法を用いた最適コドンの探索
    寺井 悟朗, 高橋 俊介, 中村 朋美, ISHII JUN, TSUGE KENJI, 浅井 潔
    日本農芸化学会2018年度大会, Mar. 2018, 名城大学 天白キャンパス

  • Enhanced protein secretion by accumulation of novel effective factors in Pichia pastoris
    ITO YOICHIRO, NAKAMURA YASUYUKI, Shimpei Aikawa, Tomohisa Hasunuma, ISHII JUN, Akihiko Kondo
    The 9th International Symposium of Innovative BioProduction Kobe (iBioK), Feb. 2018, Kobe, Japan

  • Development of G-protein-coupled receptor (GPCR)-based metabolite sensor for monitoring physiological chemical melatonin in eukaryotic yeast cells
    Takuya Tabata, NAKAMURA YASUYUKI, ISHII JUN, Akihiko Kondo
    The 9th International Symposium of Innovative BioProduction Kobe (iBioK), Feb. 2018, Kobe, Japan

  • Development of combinatorial gene assemble system and application to improvement of organic solvent tolerance in Escherichia coli
    Yoshinobu Saijo, Masahiro Tominaga, Kanae Sakai, Toshihide Matsuno, ISHII JUN, Akihiko Kondo
    The 9th International Symposium of Innovative BioProduction Kobe (iBioK), Feb. 2018, Kobe, Japan

  • A new autonomous replicating plasmid vector containing Pichia pastoris centromeric DNA
    NAKAMURA YASUYUKI, Teruyuki Nishi, Risa Noguchi, ITO YOICHIRO, Toru Watanabe, Tozo Nishiyama, Shimpei Aikawa, Tomohisa Hasunuma, ISHII JUN, Yuji Okubo, Akihiko Kondo
    The 9th International Symposium of Innovative BioProduction Kobe (iBioK), Feb. 2018, Kobe, Japan

  • Dynamic regulation of ethanol production in yeast by inactive Cas9 system
    Keisuke Morita, Fumio Matsuda, ISHII JUN, Akihiko Kondo, Hiroshi Shimizu
    2017 Asian Symposium on innovative Biorefinery in Singapore (i-BioS 2017), Dec. 2017, Nanjing, China

  • 麹菌を用いた遊離型高度不飽和脂肪酸の生産化
    玉野 孝一, Robert Sidney Cox III, Kenji Tsuge, 三浦 愛, 伊藤 あやの, Jun Ishii, 田村 具博, Akihiko Kondo, 町田 雅之
    第17回糸状菌分子生物学研究会, Nov. 2017, 糸状菌分子生物学研究会, 佐賀市立東与賀文化ホール

  • 胞子形成過程を回避した酵母の人為的有性生殖法
    福田 展雄, ISHII JUN, KONDO AKIHIKO, 本田 真也
    第68回日本生物工学会大会, Sep. 2017, 富山国際会議場

  • 長鎖DNA合成技術による高機能遺伝子デザインの可能性
    柘植 謙爾, 鈴木 武尊, 石井 純
    第69回日本生物工学会大会(ランチョンセミナー), Sep. 2017, 早稲田大学 西早稲田キャンパス

  • メラトニン濃度をモニタリングするための酵母in vivoメタボライトセンサ
    田畑 琢也, Yasuyuki Nakamura, Jun Ishii, Akihiko Kondo
    化学工学会第49回秋季大会, Sep. 2017, 化学工学会, 名古屋市

  • ピルビン酸代謝フローの転換による出芽酵母2,3-ブタンジオール生産性の向上
    森田 啓介, 松田 史生, 岡本 浩二, Jun Ishii, Akihiko Kondo, 清水 浩
    第69回日本生物工学会大会, Sep. 2017, 日本生物工学会, 早稲田大学 西早稲田キャンパス

  • Self-assembly of pentapeptide-based hydrogelators for encapsulation and release of functional compounds
    Restu Witta Kartika, Nishida Yuki, Yamamoto Shota, ISHII JUN, Maruyama Tatsuo
    第49回化学工学会秋季大会, Sep. 2017, 化学工学会, 名古屋市

  • Pichia pastorisのタンパク質分泌生産における新規有用因子の獲得とその蓄積による効果の検証
    Yoichiro Ito, Yasuyuki Nakamura, 西 輝之, 藍川 晋平, Tomohisa Hasunuma, Jun Ishii, Akihiko Kondo
    第69回日本生物工学会大会, Sep. 2017, 日本生物工学会, 早稲田大学 西早稲田キャンパス

  • Pichia pastorisにおけるセントロメア配列を含む自律複製型プラスミドの開発
    Yasuyuki Nakamura, 西 輝之, 野口 理紗, Yoichiro Ito, 渡邉 徹, 西山 陶三, 藍川 晋平, Tomohisa Hasunuma, Jun Ishii, 八十原 良彦, Akihiko Kondo
    第69回日本生物工学会大会, Sep. 2017, 日本生物工学会, 早稲田大学 西早稲田キャンパス

  • GPCR拮抗薬のポジティブ検出を可能とする反転型レポーター発現系
    福田 展雄, ISHII JUN, 本田 真也
    第69回日本生物工学会大会, Sep. 2017, 早稲田大学 西早稲田キャンパス

  • 特定の生理活性物質を濃度依存的に感知する酵母メタボライトセンサの開発
    田畑 琢也, 中村 泰之, ISHII JUN, KONDO AKIHIKO
    生物工学若手研究者の集い 夏のセミナー2017, Jul. 2017, 日本生物工学会 若手会, ツネイシしまなみビレッジ

  • 酵母代謝改変のための遺伝子のノックダウン/ノックアウト法の開発
    光増 遼太郎, 冨永 将大, ISHII JUN, KONDO AKIHIKO
    生物工学若手研究者の集い 夏のセミナー2017, Jul. 2017, 日本生物工学会 若手会, ツネイシしまなみビレッジ

  • 酵母遺伝子スイッチの進化工学のためのON/OFF選抜法の開発
    能崎 健太, 冨永 将大, 河合(野間) 繁子, 梅野 太輔, ISHII JUN, KONDO AKIHIKO
    生物工学若手研究者の集い 夏のセミナー2017, Jul. 2017, 日本生物工学会 若手会, ツネイシしまなみビレッジ

  • 酵母シグナル伝達を利用したGγ recruitment systemによるタンパク質-ペプチド間相互作用の検出と変異体スクリーニング
    松原 大希, 海嶋 美里, ISHII JUN, KONDO AKIHIKO
    生物工学若手研究者の集い 夏のセミナー2017, Jul. 2017, 日本生物工学会 若手会, ツネイシしまなみビレッジ

  • 油性酵母Lipomyces starkeyiを利用したテルペン生産系の開発
    金本 浩介, 小島 基, ISHII JUN, KONDO AKIHIKO, 三沢 典彦, 播本 孝史
    第11回日本ゲノム微生物学会年会, Mar. 2017, 日本ゲノム微生物学会, 慶應義塾大学湘南藤沢キャンパス

  • 油性酵母 (Lipomyces starkeyi) によるアルテミシニン前駆体の生産
    小島 基, 鈴木 宗典, 金本 浩介, Jun Ishii, Akihiko, 播本 孝史
    日本農芸化学会2017年度大会, Mar. 2017, 日本農芸化学会, 京都女子大学

  • 酵母を用いたHMG-CoA reductase阻害剤FR901512生合成遺伝子クラスター解析と新規物質生産の可能性
    伊東 広哉, 宮村 由紀, 竹田 至, 松井 真, ISHII JUN, 熊谷 俊高, 町田 雅之, 有田 正規, 柴田 孝
    第11回日本ゲノム微生物学会年会, Mar. 2017, 日本ゲノム微生物学会, 慶應義塾大学湘南藤沢キャンパス

  • 酵母でのモノづくり細胞のエンジニアリング
    ISHII JUN
    第11回日本ゲノム微生物学会年会, Mar. 2017, 日本ゲノム微生物学会, 慶應義塾大学湘南藤沢キャンパス

  • メラトニン濃度をモニタリングするための酵母in vivoバイオセンサー
    田畑 琢也, 中村 泰之, Ishii Jun, KONDO Akihiko
    第19回化学工学会学生発表会, Mar. 2017, 化学工学会関西支部, 豊中市

  • マイトファジー抑制による出芽酵母の細胞質物質生産能力向上
    森田 啓介, 松田 史生, 岡本 浩二, Jun Ishii, Akihiko, 清水 浩
    日本農芸化学会2017年度大会, Mar. 2017, 日本農芸化学会, 京都女子大学

  • In vitro selection of novel peptide agonists for human somatostatin receptor subtype-2 using a water-in-oil microdroplet platform
    T. Sakurai, R. Iizuka, ISHII JUN, NAKAMURA YASUYUKI, KONDO AKIHIKO, A. Iguchi, D.H. Yoon, T. Sekiguchi, S. Shoji, T. Funatsu
    The 20th International Conference on Miniaturized Systems for Chemistry and Life Sciences (µTAS 2016), Oct. 2016, Dublin, Ireland

  • イソブタノール生産酵母の構築:オルガネラ局在の検討
    森田 啓介, 松田 史生, 伊田 賢吾, ISHII JUN, KONDO Akihiko, 清水 浩
    第68回日本生物工学会大会, Sep. 2016, 日本生物工学会, 富山国際会議場

  • Metabolic and cell surface engineering to produce isobutanol from lignocellulosic baiomass in yeast Saccharomyces cerevisiae
    Ishii Jun, Fumio Matsuda, Shuhei Ishikado, Kengo Ida, KONDO Akihiko
    The 14th International Congress on Yeasts (ICY2016), Sep. 2016, ICY14組織委員会, Awaji, Japan

  • Enhanced protein secretion by accumulation of novel effective factors in Pichia pastoris
    Yoichiro Ito, Yasuyuki Nakamura, Aikawa Shimpei, Hasunuma Tomohisa, Ishii Jun, KONDO Akihiko
    The 14th International Congress on Yeasts (ICY2016), Sep. 2016, ICY14組織委員会, 淡路市

  • Engineering of mitochondrial isobutanol production in Saccharomyces cerevisiae
    Keisuke Morita, Fumio Matsuda, Koji Okamoto, Kengo Ida, ISHII JUN, Akihiko Kondo, Hiroshi Shimizu
    The 14th International Congress on Yeasts (ICY2016), Sep. 2016, Awaji, Japan

  • A new autonomous replicating plasmid vector containing centromere DNA sequence of Pichia pastoris
    NAKAMURA YASUYUKI, Teruyuki Nishi, Risa Noguchi, ITO YOICHIRO, Toru Watanabe, Tozo Nishiyama, Shimpei Aikawa, Tomohisa Hasunuma, ISHII JUN, Yoshihiko Yasohara, Akihiko Kondo
    The 14th International Congress on Yeasts (ICY2016), Sep. 2016, Awaji, Japan

  • 微生物バイオによるテルペン系化合物の生産法開発
    金本 浩介, 竹村 秀史, 原田 尚志, 三沢 典彦, ISHII JUN, KONDO AKIHIKO, 播本 孝史
    第6回植物生理化学会シンポジウム, Jul. 2016, 大阪府立大学中百舌鳥キャンパス

  • 培地中のグルコース濃度をモニタリングできる酵母in vivo バイオセンサの開発
    橋弘樹, 中村泰之, ISHII JUN, KONDO Akihiko
    生物工学若手研究者の集い 夏のセミナー2016, Jul. 2016, 生物工学会, 府中市

  • 出芽酵母における酵素連結法を用いた代謝フラックス改変
    濱田省吾, Nishimura Yuuya, ISHII JUN, KONDO Akihiko
    生物工学若手研究者の集い 夏のセミナー2016, Jul. 2016, 生物工学会, 府中市

  • メラトニン濃度をモニタリングするための酵母in vivoバイオセンサの開発
    田畑琢也, 中村泰之, ISHII JUN, KONDO Akihiko
    生物工学若手研究者の集い 夏のセミナー2016, Jul. 2016, 生物工学会, 府中市

  • Metabolic and cell surface engineering to produce isobutanol from lignocellulosic biomass in yeast Saccharomyces cerevisiae
    Ishii Jun, Fumio Matsuda, Shuhei Ishikado, Kengo Ida, KONDO Akihiko
    Metabolic Engineering 11, Jun. 2016, Metabolic Engineering 11, Awaji, Japan

  • Heterologous gene expression in Saccharomyces cerevisiae for higher isobutanol production
    Keisuke Morita, Fumio Matsuda, Ishii Jun, KONDO Akihiko, Hiroshi Shimizu
    Metabolic Engineering 11, Jun. 2016, Metabolic Engineering 11, 淡路市

  • Expression of varied GFPs in Saccharomyces cerevisiae: comparison of codon-optimized and non-codon-optimized GFPs
    Toshihide Matsuno, Misato Kaishima, Ishii Jun, KONDO Akihiko
    Metabolic Engineering 11, Jun. 2016, Metabolic Engineering 11, 淡路市

  • Development of selection method for directed evolution of genetic switches in Saccharomyces cerevisiae
    Masahiro Tominaga, Shigeko Kawai-Noma, Daisuke Umeno, Ishii Jun, KONDO Akihiko
    Metabolic Engineering 11, Jun. 2016, Metabolic Engineering 11, 淡路市

  • Development of Platform Yeast Strain Capable of Direct Fermentation of Raw Biomass to Ethanol
    PRIHARDI KAHAR, Akiho Itomi, Ahmad Thontowi, Hiromi Otsuka, Chie Kihira, Jaemin Lee, Ario Betha Juanssil fero, Apridah Cameliawati Djohan, Yopi Sunarya, Ishii Jun, OGINO Chiaki, Banbang Prasetya, KONDO Akihiko
    Metabolic Engineering 11, Jun. 2016, Metabolic Engineering 11, 淡路市

  • 1-Propanol production of S. cerevisiae engineering 2-ketobutyrate biosynthetic pathway
    Nishimura Yuuya, Ishii Jun, KONDO Akihiko
    Metabolic Engineering 11, Jun. 2016, Metabolic Engineering 11, 淡路市

  • 大腸菌における1,4-ブタンジオール人工生合成経路構築
    Eguchi Kouichi, Kudou Motonori, Araki, Ishii Jun, Yamamoto Hiroaki, Kondo Akihiko
    日本農芸化学会2016年度大会, Mar. 2016

  • 異種遺伝子発現による出芽酵母イソブタノール合成活性化
    Morita Keisuke, Matsuda Fumio, Ishii Jun, Kondo Akihiko, Shimizu Hirosi
    日本農芸化学会2016年度大会, Mar. 2016

  • マンナンバイオマスからのエタノール生産:βーマンナナーゼとβーマンノシダーゼを細胞表層に提示した出芽酵母の開発
    ISHII JUN, OKAZAKI FUMIYOSHI, Apridah Cameliawati Djohan, HARA KIYOTAKA, ASAI NANAMI, Ade Andriani, TERAMURA HIROSHI, Prihardi Kahar, YOPI, Bambang Prasetya, OGINO CHIAKI, AKIHIKO KONDO
    日本農芸化学会2016年度大会, Mar. 2016

  • パスウェイエンジニアリングを用いたフリージア・エアリーパープル及びエアリーピーチの新規セキステルペン合成酵素遺伝子の機能解析
    Higuchi Yuki, Hattan Junichirou, Ohono Fumina, Ito Tomoko, Sibuya Yurika, Watanabe Arisa, Ishii Jun, Kondo Akihiko, Misawa Norihiko
    日本農芸化学会2016年度大会, Mar. 2016

  • タキサジエンの10位を水酸化する新規シトクロムP450 遺伝子の単離と機能解析
    Komori Aya, Takemura Hidefumi, Kubo Takashi, Ishii Jun, Machida Masayuki, Kondo Akihiko, Harimoto Takashi, Suzuki Munenori
    日本農芸化学会2016年度大会, Mar. 2016

  • From mannan and lignocellulosic biomass to biochemicals: cell surface display and metabolic engineering in yeast Saccharomyces cerevisiae
    ISHII JUN, OKAZAKI FUMIYOSHI, Shuhei Ishikado, Apridah Cameliawati Djohan, Nanami Asai-Nakashima, Yopi, Fumio Matsuda, OGINO CHIAKI, AKIHIKO KONDO
    The 6th iBioK Asian Workshop, Mar. 2016, Kobe

  • 2,3-ブタンジオール高生産出芽酵母株の構築
    Matsuda Fumio, Ida Kengo, Kinoshita Syohei, Morita Keisuke, Ishii Hiroko, Tomita Atsumi, Ishii Jun, Kondo Akihiko
    日本農芸化学会2016年度大会, Mar. 2016

  • Specific drug delivery for target cancer tumor using affibody-displaying bionanocapsule/liposome complex
    Yuya Nishimura, Jun Ishii, Chiaki Ogino, Akihiko Kondo
    The 2015 International Chemical Congress of Pacific Basin Societies (Pacifichem 2015), Dec. 2015

  • Live cell imaging and membrane protein mapping with atomic force microscope
    Musashi Takenaka, T. Kobayashi, Yusuke Miyachi, Kentarou Inokuma, Jun Ishii, Tomohisa Hasunuma, Chiaki Ogino, Akihiko Kondo
    The 2015 International Chemical Congress of Pacific Basin Societies (Pacifichem 2015), Dec. 2015

  • High-resolution, quantitative analysis for measuring heterogeneities of G-protein signaling at single-cell levels in yeast
    Jun Ishii, Akihiko Kondo
    The 2015 International Chemical Congress of Pacific Basin Societies (Pacifichem 2015), Dec. 2015

  • Desired alteration of protein affinities using competitive screening system
    Misato Kaishima, Nobuo Fukuda, Jun Ishii, Akihiko Kondo
    The 2015 International Chemical Congress of Pacific Basin Societies (Pacifichem 2015), Dec. 2015

  • 酵母シグナル伝達を利用した結合タンパク質の選択的スクリーニング
    Matsubara Taiki, Kaishima Misato, Ishii Jun, Kondo Akihiko
    33th Yeast Workshop, Nov. 2015

  • 酵母シグナル伝達を用いた標的膜タンパク質に対するバイオメディカル分子スクリーニングシステム
    海嶋 美里, 福田 展雄, Ishii jun, Kondo Akihiko
    33th Yeast Workshop, Nov. 2015

  • ヒトニューロテンシン受容体のリガンド検出のための酵母バイオセンサーの開発
    橋 弘樹, Nakamura Yasuyuki, Kondo Akihiko, Ishii Jun
    33th Yeast Workshop, Nov. 2015

  • 大腸菌パスウェイエンジニアリングによるパクリタキセル前駆体タキサジエン酸化物の生産
    Takemura Hidefumi, Suzuki Munenori, Harada Hisasi, Misawa Norihiko, Ishii Jun, Kondou Akihiko, 播本 孝史
    第67回日本生物工学会大会 国際シンポジウム, Oct. 2015

  • フリージア・エアリーパープルから新規セスキテルペン合成酵素遺伝子の単離と機能解析
    Higuchi Yuuki, Hattan Jyunichirou, Oono Fumina, Itou Tomoko, Shibutani Yurika, Watanabe Arisa, Ishii Jyun, Kondou Akihiko, Shindou Kazutoshi, Misawa Norihiko
    第67回日本生物工学会大会 国際シンポジウム, Oct. 2015

  • ヒトニューロテンシン受容体におけるリガンド探索のための酵母バイオセンサー
    Hashi Hiroki, Nakamura Yasuyuki, Ishii Jun, Kondou Akihiko
    第67回日本生物工学会大会 国際シンポジウム, Oct. 2015

  • カンツバキから新規セスキテルペン合成酵素遺伝子の単離と機能解析
    Hattan Jyunichirou, Shindou Kazutoshi, Oono Shina, Higuchi Yuuki, Ishi Jun, Kondo Akihiko, Misawa Norihiko
    第67回日本生物工学会大会 国際シンポジウム, Oct. 2015

  • 合成生物学的アプローチを目指した酵母プロティンタグの開発
    Nishimura Yuya, Ishii Jun, Kondo Akihiko
    化学工学会第47回秋季大会, Sep. 2015

  • 長期継代培養によるエタノール非生産酵母の増殖改善株の取得
    森田 啓介, 富田 淳美, 松田 史生, ISHII JUN, KONDO AKIHIKO, 清水 浩
    日本農芸化学会2015年度大会, Mar. 2015, 日本農芸化学会, 岡山市

  • 酵母における1-propanol発酵生産
    西村 勇哉, ISHII JUN, KONDO AKIHIKO
    化学工学会第80年会, Mar. 2015, 化学工学会, 江東区

  • ヒトニューロテンシン受容体のリガンド探索のための酵母1細胞分析用バイオセンサー
    橋 弘樹, 中村 泰之, ISHII JUN, KONDO AKIHIKO
    第17回化学工学会学生発表会, Mar. 2015, 化学工学会, 徳島市

  • 酵母における合成生物工学ツールの開発
    ISHII JUN
    合成生物工学シンポジウム, Nov. 2014, 文部科学省先端融合領域イノベーション創出拠点形成プログラム 「バイオプロダクション次世代農工連携拠点 」, 神戸大学 統合研究拠点 コンベンションホール

  • G 蛋白質共役型受容体の二量体形成およびシグナル伝達同時解析のための二色蛍光レポーターによる酵母細胞設計
    ISHII JUN, 中村 泰之, 竹本 紀加, KONDO AKIHIKO
    「細胞を創る」研究会7.0, Nov. 2014, 「細胞を創る」研究会, 東京大学 弥生講堂一条ホール

  • Peptide-based ligand screening system for G protein-coupled receptors using water-in-oil microdroplets
    Takashi Sakurai, Ryo Iizuka, Rui Sekine, Yasuyuki Nakamura, Dong Hyun Yoon, Tetsushi Sekiguchi, Jun Ishii, Akihiko Kondo, Shuichi Shoji, Takashi Funatsu
    The 18th International Conference on Miniaturized Systems for Chemistry and Life Sciences (µTAS 2014), Oct. 2014, San Antonio, TX, USA

  • Genetic engineering for altering metabolic flow in Saccharomyces cerevisiae to produce higher alcohols
    ISHII JUN, Fumio Matsuda, AKIHIKO KONDO
    The 31st International Specialised Symposium on Yeast (ISSY 31), Oct. 2014, Vipava and Nova Gorica, Slovenia

  • 二次代謝産物を指標とした有用油性酵母のクラスター解析
    小島 基, 鈴木 宗典, 原田 尚志, 三沢 典彦, ISHII JUN, KONDO AKIHIKO, 播本 孝史
    第66回日本生物工学会大会, Sep. 2014, 日本生物工学会, 札幌市

  • 大腸菌のパスウェイエンジニアリングによる安価な基質からのパクリタキセル前駆体タキサジエンの効率的生産
    竹村 秀史, 鈴木 宗典, 梅野 太輔, 原田 尚志, 三沢 典彦, ISHII JUN, KONDO AKIHIKO, 播本 孝史
    第66回日本生物工学会大会, Sep. 2014, 日本生物工学会, 札幌市

  • 出芽酵母における分割ルシフェラーゼを利用したGPCR リガンド応答検出法の応用
    浅川 賢史, 福谷 洋介, ISHII JUN, KONDO AKIHIKO, 小澤 岳晶, 養王田 正文
    第66回日本生物工学会大会, Sep. 2014, 日本生物工学会, 札幌市

  • 出芽酵母 Saccharomyces cerevisiae での嗅覚受容体匂い分子応答における匂い結合タンパク質の効果
    堀 綾佳, 福谷 洋介, 塚田 聡史, 佐藤 令一, ISHII JUN, KONDO AKIHIKO, 松波 宏明, 養王田 正文
    第66回日本生物工学会大会, Sep. 2014, 日本生物工学会, 札幌市

  • pH 応答性ペプチドGALA を表層提示したバイオナノカプセルのエンドソーム脱出
    西村 勇哉, 江澤 僚将, ISHII JUN, OGINO CHIAKI, KONDO AKIHIKO
    第66回日本生物工学会大会, Sep. 2014, 日本生物工学会, 札幌市

  • G タンパク質共役型受容体の二量体形成検出のためのゲノム編集技術の開発
    橋本 貴理, 中村 泰之, ISHII JUN, KONDO AKIHIKO
    第66回日本生物工学会大会, Sep. 2014, 日本生物工学会, 札幌市

  • pH応答性膜融合ペプチドGALAを表層提示したバイオナノカプセルのエンドソーム脱出
    西村 勇哉, 竹田 光一, 江澤 僚将, ISHII JUN, OGINO CHIAKI, KONDO AKIHIKO
    生物工学若手研究者の集い夏のセミナー, Jul. 2014, 生物工学会, 神戸市

  • 色素生産を指標としたCoA代謝経路の評価系
    磯貝 章太, ISHII JUN, HASUNUMA TOMOHISA, KONDO AKIHIKO
    生物工学若手研究者の集い夏のセミナー, Jul. 2014, 生物工学会, 神戸市

  • 上皮成長因子受容体を特異的に認識するAffibody提示バイオナノカプセルの開発
    江澤 僚将, 西村 勇哉, ISHII JUN, OGINO CHIAKI, KONDO AKIHIKO
    生物工学若手研究者の集い夏のセミナー, Jul. 2014, 生物工学会, 神戸市

  • 酵母シグナル伝達機構を基盤としたアフィニティ改変タンパク質スクリーニングシステムの開発
    海嶋 美里, 福田 展雄, ISHII JUN, KONDO AKIHIKO
    生物工学若手研究者の集い夏のセミナー, Jul. 2014, 生物工学会, 神戸市

  • アンジオテンシン受容体のリガンド探索のための酵母バイオセンサーの開発
    中村 泰之, ISHII JUN, KONDO AKIHIKO
    生物工学若手研究者の集い夏のセミナー, Jul. 2014, 生物工学会, 神戸市

  • G蛋白質共役型受容体の二量体形成に応答して機能するゲノム編集技術の開発
    橋本 貴理, 中村 泰之, ISHII JUN, KONDO AKIHIKO
    生物工学若手研究者の集い夏のセミナー, Jul. 2014, 生物工学会, 神戸市

  • Metabolic engineering of yeast central metabolism for higher alcohol production
    Fumio Matsuda, Jun Ishii, Syunsuke Nishino, Keisuke Morita, Akihiko Kondo, Hiroshi Shimizu
    Metabolic Engineering X, Jun. 2014, Society for Biological Engineering, Vancouver, Canada

  • Genetic engineering to produce higher alcohols in yeast Saccharomyces cerevisiae
    Jun Ishii, Fumio Matsuda, Kengo Ida, Akihiko Kondo
    Metabolic Engineering X, Jun. 2014, Society for Biological Engineering, Vancouver, Canada

  • 代謝シミュレーションにもとづく酵母イソブタノール生産能の向上
    松田 史生, ISHII JUN, 近藤 貴志, 伊田 賢吾, 手塚 裕紀, KONDO AKIHIKO
    日本農芸化学会2014年度大会, Mar. 2014, 日本農芸化学会, 川崎市

  • ED経路導入による酵母中央代謝経路の拡張
    森田 啓介, 梶畠 秀一, ISHII JUN, 松田 史生, 清水 浩
    日本農芸化学会2014年度大会, Mar. 2014, 日本農芸化学会, 川崎市

  • 酵母の遺伝情報を操作する
    ISHII JUN
    「細胞を創る」研究会6.0, Nov. 2013, 「細胞を創る」研究会, 鶴岡メタボロームキャンパス レクチャーホール

  • Metabolic engineering of yeast Saccharomyces cerevisiae for increased isobutanol production
    ISHII JUN, Fumio Matsuda, AKIHIKO KONDO
    2013 Asian Symposium on innovative Biorefinery in Beijing (i-BioB 2013), Oct. 2013, Beijing, China

  • 上皮成長因子受容体を特異的に認識するAffibody提示バイオナノカプセルの開発
    江澤 僚将, 西村 勇哉, ISHII JUN, OGINO CHIAKI, KONDO AKIHIKO
    第65回日本生物工学会大会, Sep. 2013, 日本生物工学会, 広島市

  • 出芽酵母Saccharomyces cerevisiaeでの嗅覚受容体の機能的発現におけるアクセサリータンパク質の効果
    堀 綾佳, 福谷 洋介, ISHII JUN, KONDO AKIHIKO, 松波 宏明, 養王田 正文
    第65回日本生物工学会大会, Sep. 2013, 日本生物工学会, 広島市

  • 酵母蛍光レポーターの改変によるヒト由来7回膜受容体の高感度リガンド検出システム
    中村 泰之, ISHII JUN, KONDO AKIHIKO
    第65回日本生物工学会大会, Sep. 2013, 日本生物工学会, 広島市

  • ヒト受容体リガンド探索のための酵母蛍光レポーター高感度アッセイシステムの開発とその応用
    中村 泰之, ISHII JUN, KONDO AKIHIKO
    化学工学会第45回秋季大会, Sep. 2013, 化学工学会, 岡山市

  • イソブタノール生産酵母におけるトランスヒドロゲナーゼ様シャントの活性化
    ISHII JUN, 松田 史生, 近藤 貴志, KONDO AKIHIKO
    第65回日本生物工学会大会, Sep. 2013, 日本生物工学会, 広島市

  • G蛋白質共役型受容体の二量体形成およびシグナル伝達の同時解析システム
    竹本 紀加, 中村 泰之, ISHII JUN, KONDO AKIHIKO
    第65回日本生物工学会大会, Sep. 2013, 日本生物工学会, 広島市

  • Metabolic pathway engineering of yeast Saccharomyces cerevisiae for isobutanol production
    Jun Ishii, Fumio Matsuda, Akihiko Kondo
    The 26th International Conference on Yeast Genetics and Molecular Biology (Yeast2013), Aug. 2013, Committee of the 26th International Conference on Yeast Genetics and Molecular Biology, Frankfurt am Main, Germany

  • A yeast-based simultaneous method to analyze dimerization and signaling of G-protein-coupled receptor by dual-color reporter
    Yasuyuki Nakamura, Norika Takemoto, Jun Ishii, Akihiko Kondo
    The 19th Symposium of Young Asian Biochemical Engineers' Community (YABEC2013), Aug. 2013, 生物工学会, Xinjiang,China

  • A selection system exploiting yeast signal transduction machinery to create desirably affinity-altered protein variants.
    Misato Kaishima, Nobuo Fukuda, Jun Ishii, Akihiko Kondo
    The 26th International Conference on Yeast Genetics and Molecular Biology (Yeast2013), Aug. 2013, Committee of the 26th International Conference on Yeast Genetics and Molecular Biology, Frankfurt am Main, Germany

  • A selection method of desirably affinity-altered protein mutants using cellular signaling in living yeast cells
    Misato Kaishima, Nobuo Fukuda, Jun Ishii, Akihiko Kondo
    The 19th Symposium of Young Asian Biochemical Engineers' Community (YABEC2013), Aug. 2013, 生物工学会, Xinjiang,China

  • ヒト由来7回膜受容体の酵母蛍光レポーター改変型高感度リガンド検出システムの開発とその応用
    中村 泰之, ISHII JUN, KONDO AKIHIKO
    生物工学若手研究者の集い 夏のセミナー2013, Jul. 2013, 生物工学会, 宮崎市

  • Saccharomyces cerevisiaeにおけるイソブタノール高生産のための代謝改変
    ISHII JUN, 松田 史生, 近藤 貴志, KONDO AKIHIKO
    生物工学若手研究者の集い 夏のセミナー2013, Jul. 2013, 生物工学会, 宮崎市

  • Metabolically engineered yeast Saccharomyces cerevisiae for increased isobutanol production
    Jun Ishii, Fumio Matsuda, Akihiko Kondo
    World Biotechnology Congress 2013 (WBC2013), Jun. 2013, Eureka Conferences, Boston, USA

  • Developing a Screening Method for Desirably Altering Affinities of Protein Mutants Based on Yeast Cellular Signaling
    Misato Kaishima, Nobuo Fukuda, Jun Ishii, Akihiko Kondo
    第35回日本分子生物学会年会, Dec. 2012, 日本分子生物学会, 福岡市

  • Interaction survey between GPCR and ligand on the surface of yeast by AFM
    Musashi Takenaka, Tomokazu Amino, Yusuke Miyachi, Jun Ishii, Chiaki Ogino, Akihiko Kondo
    International Joint Symposium on Single-Cell Analysis (The 6th International Workshop on Approaches to Single-Cell Analysis & The 8th International Forum on Post-Genome Technologies), Nov. 2012, The Society for Single-Cell Surveyor, 京都市

  • Improvement of reporter sensitivity for human G-protein-coupled receptor signaling assay in yeast cells by flow cytometry
    Yasuyuki Nakamura, Jun Ishii, Akihiko Kondo
    International Joint Symposium on Single-Cell Analysis (The 6th International Workshop on Approaches to Single-Cell Analysis & The 8th International Forum on Post-Genome Technologies), Nov. 2012, The Society for Single-Cell Surveyor, 京都市

  • A technique to screen heterdimerizable partners against target human G-protein-coupled receptors in living yeast cells
    Jun Ishii, Yasuyuki Nakamura, Akihiko Kondo
    International Joint Symposium on Single-Cell Analysis (The 6th International Workshop on Approaches to Single-Cell Analysis & The 8th International Forum on Post-Genome Technologies), Nov. 2012, The Society for Single-Cell Surveyor, 京都市

  • A selection method of desirably affinity-altered protein mutants using cellular signaling in living yeast cells
    Misato Kaishima, Nobuo Fukuda, Jun Ishii, Akihiko Kondo
    International Joint Symposium on Single-Cell Analysis (The 6th International Workshop on Approaches to Single-Cell Analysis & The 8th International Forum on Post-Genome Technologies), Nov. 2012, The Society for Single-Cell Surveyor, 京都市

  • 出芽酵母を用いた嗅覚受容体リガンドアッセイシステムの最適化
    福谷 洋介, ISHII JUN, 野口 恵一, KONDO AKIHIKO
    第64回日本生物工学会大会, Oct. 2012, 日本生物工学会, 神戸市

  • Yeast-based ligand assay system for detecting G protein-coupled receptor activation in water-in-oil droplets
    Takashi Sakurai, Ryo Iizuka, Yasuyuki Tanigaki, Rui Sekine, Dong Hyun Yoon, Tetsushi Sekiguchi, ISHII JUN, KONDO AKIHIKO, Naoto Nemoto, Shuichi Shoji, Takashi Funatsu
    The 16th International Conference on Miniaturized Systems for Chemistry and Life Sciences (µTAS 2012), Oct. 2012, Okinawa, Japan

  • Genetic engineering of valine biosynthesis for isobutanol production in Saccharomyces cerevisiae
    Jun Ishii, Fumio Matsuda, Kengo Ida, Hironori Tezuka, Takashi Kondo, Chiaki Ogino, Akihiko Kondo
    The 18th Symposium of Young Asian Biochemical Engineers' Community (YABEC2012), Oct. 2012, Society for Biological Engineering, Tokushima, Japan

  • A yeast-based screening method to identify hetero-dimerizable partners against target human G-protein-coupled receptor
    Yasuyuki Nakamura, Jun Ishii, Akihiko Kondo
    The 18th Symposium of Young Asian Biochemical Engineers' Community (YABEC2012), Oct. 2012, Society for Biological Engineering, Tokushima, Japan

  • A new technology to screen desirably affinity-altered proteins based on yeast signal transduction machinery
    Misato Kaishima, Nobuo Fukuda, Jun Ishii, Akihiko Kondo
    The 18th Symposium of Young Asian Biochemical Engineers' Community (YABEC2012), Oct. 2012, Society for Biological Engineering, Tokushima, Japan

  • 非天然型経路を導入した新規イソブタノール生産酵母の開発
    伊田 賢吾, MATSUDA FUMIO, 近藤 貴志, ISHII JUN, KONDO AKIHIKO
    日本農芸化学会関西支部大会, Sep. 2012, 日本農芸化学会, 亀岡市

  • Complex carriers of affibody-displaying bio-nanocapsule and composition-varied liposomes for HER2-expressing breast cancer cell-specific protein delivery
    Yuya Nishimura, Jun Ishii, Fumiyoshi Okazaki, Chiaki Ogino, Akihiko Kondo
    The 15th International Biotechnology Symposium (IBS 2012), Sep. 2012, The Korean Society for Biotechnology and Bioengineering, Deague, Korea

  • Genetically enhanced valine biosynthesis and the Ehrlich pathway for isobutanol production in Saccharomyces cerevisiae
    Jun Ishii, Takashi Kondo, Hironori Tezuka, Kengo Ida, Fumio Matsuda, Akihiko Kondo
    The 13th International Congress on Yeasts (ICY2012), Aug. 2012, ICY 2012 Organizing Committee, Madison, WI, USA

  • Genetic engineering to enhance the Ehrlich pathway and alter carbon flux for increased isobutanol production by Saccharomyces cerevisiae
    Jun Ishii, Takashi Kondo, Hiroyuki Tezuka, Fumio Matsuda, Chiaki Ogino, Akihiko Kondo
    Metabolic EngineeringⅨ, Jun. 2012, Engineering Conferences International, Biarritz, France

  • Effect of metabolic inhibitors on yeast central metabolism
    MATSUDA Fumio, Ishii Jun, Kondo Akihiko
    Metabolic EngineeringⅨ, Jun. 2012, Engineering Conferences International, Biarritz, France

  • Combinatorial screen of human G-protein-coupled receptor heterodimer pairs by using a split-ubiquitin yeast two-hybrid system
    Yasuyuki Nakamura, Jun Ishii, Akihiko Kondo
    Japan-Finland Biotechnology Symposium 2012, Jun. 2012, Sendai, Japan

  • A new technology to screen desirably affinity-altered proteins
    Misato Kaishima, Nobuo Fukuda, Jun Ishii, Akihiko Kondo
    Japan-Finland Biotechnology Symposium 2012, Jun. 2012, Sendai, Japan

  • Saccharomyces cerevisiaeにおけるイソブタノール生産性向上のためのEhrlich経路強化と炭素フラックス改変戦略
    手塚 裕紀, 近藤 貴志, 石井 純
    日本農芸化学会2012年度大会, Mar. 2012, 日本農芸化学会, 京都市

  • Amplification and smoothing of signal activation from human G-protein-coupled receptor for enriching agonist stimulation in yeast
    小田 麻美, 石井 純, 福田 展雄, 藤原 俊伸, 荻野 千秋, 近藤 昭彦
    第34回日本分子生物学会年会, Dec. 2011, 日本分子生物学会, 横浜市

  • A yeast two-hybrid method to identify heterodimerization pairs of human G protein-coupled receptors
    Yasuyuki Nakamura, Jun Ishii, Akihiko Kondo
    The 17th Symposium of Young Asian Biochemical Engineers' Community (YABEC2011), Oct. 2011, Society for Biological Engineering, Incheon, Korea

  • 次世代バイオアルコール生産酵母の代謝デザイン
    松田 史生, 古澤 力, 石井 純
    第63回日本生物工学会大会, Sep. 2011, 日本生物工学会, 小金井市

  • マウス嗅覚受容体の出芽酵母での機能的発現における N 末端およびC 末端配列置換の効果
    福谷 洋介, 中村 智子, 萬 麻衣子, 野口 恵一, 石井 純
    第63回日本生物工学会大会, Sep. 2011, 日本生物工学会, 小金井市

  • フラン化合物存在下でのXR-XDH 発現酵母を用いたキシロース発酵におけるNADH 依存性Adh1過剰発現のインパクト
    石井 純, 吉村 一也, 蓮沼 誠久, 近藤 昭彦
    第63回日本生物工学会大会, Sep. 2011, 日本生物工学会, 小金井市

  • バイオマス資源からの BIO-1,3-Butanediol 発酵生産
    岡林 智仁, 中島 賢則, 山本 浩明, 近藤 貴志, 石井 純
    第63回日本生物工学会大会, Sep. 2011, 日本生物工学会, 小金井市

  • タンパク質アファニティを自在に改変するための新規スクリーニングシステムの開発
    海嶋 美里, 福田 展雄, 石井 純
    第63回日本生物工学会大会, Sep. 2011, 日本生物工学会, 小金井市

  • G 蛋白質共役型受容体のヘテロダイマー形成ペアの探索のための新規検出システムの開発
    中村 泰之, 石井 純
    第63回日本生物工学会大会, Sep. 2011, 日本生物工学会, 小金井市

  • Affibody 提示Bio-nanocapsule を用いたER2 発現癌細胞へのタンパク質送達システム
    西村 勇哉, 三村 和起子, 竹田 光一, 石井 純, 岡崎 文美, 荻野 千秋, 近藤 昭彦
    第63回日本生物工学会大会, Sep. 2011, 日本生物工学会, 小金井市

  • 代謝工学を活用したイソブタノール生産酵母の創製
    手塚 裕紀, KONDO Takashi, ISHII Jun, OGINO Chiaki, KONDO Akihiko
    化学工学会第76年会, Mar. 2011, 日本化学会, 東京都 小金井市

  • 酵母GPCRシグナル伝達系を用いたタンパク質アフィニティ改変技術の開発
    海嶋 美里, 福田 展雄, ISHII Jun, KONDO Akihiko
    化学工学会第15回学生発表会, Mar. 2011, (社)化学工学会, 神戸市

  • バイオマス資源からの BIO-1,3-Butanediol 発酵生産-2 -グルコースから 1,3-Butanediol の合成-
    中島 賢則, 岡林 智仁, 山本 浩明, KONDO Takashi, ISHII Jun, KONDO Akihiko
    日本農芸化学会2011年度大会, Mar. 2011, 日本農芸化学会, 京都市

  • バイオマス資源からの BIO-1,3-Butanediol 発酵生産-1
    岡林 智仁, 中島 賢則, 山本 浩明, KONDO Takashi, ISHII Jun, KONDO Akihiko
    日本農芸化学会2011年度大会, Mar. 2011, 日本農芸化学会, 京都市

  • Single-cell analysis revealed correlation between yeast GPCR signaling and plasmid retention
    Jun Ishii, Chiaki Ogino, Akihiko Kondo
    The 5th International Workshop on Approaches to Single-Cell Analysis, Mar. 2011, Tokyo, Japan

  • Expression and functional characterization of olfactory receptors in Saccharomyces cerevisiae
    Yosuke Fukutani, Jun Ishii, Keiichi Noguchi, Noriyuki Ishii, Akihiko Kondo, Masafumi Yohda
    The 5th International Workshop on Approaches to Single-Cell Analysis, Mar. 2011, Tokyo, Japan

  • AFMを用いた生細胞表層のレセプター・リガンド間相互作用解析と細胞応答の観察
    OGINO Chiaki, 野坂 和輝, ISHII Jun, 宮地 佑典, KONDO Akihiko
    日本化学会第91春季年会, Mar. 2011, 化学工学会, 横浜市

  • High-resolution, quantitative signalling assay for G-protein-coupled receptors by flow cytometry
    Akihiko Kondo, Jun Ishii, Nobuo Fukuda, Chiaki Ogino
    The 2010 International Chemical Congress of Pacific Basin Societies (Pacifichem 2010), Dec. 2010, Honolulu, Hawaii, USA

  • ユビキチン分割体を用いたG 蛋白質共役型受容体二量体化検出システムとドメイン解析への応用
    中村 泰之, ISHII Jun, KONDO Akihiko
    第62回日本生物工学会大会, Oct. 2010, (社)日本生物工学会, 宮崎市

  • キシロース資化性Saccharomyces cerevisiae におけるアルコールデヒドロゲナーゼ過剰発現およびフラン化合物存在下でのキシロース発酵特性
    吉村 一也, ISHII Jun, HASUNUMA Tomohisa, KONDO Akihiko
    第62回日本生物工学会大会, Oct. 2010, (社)日本生物工学会, 宮崎市

  • 1細胞アッセイ技術に基づくGPCR 解析、アゴニスト探索法の開発
    KONDO Akihiko, ISHII Jun, OGINO Chiaki
    第62回日本生物工学会大会, Oct. 2010, (社)日本生物工学会, 宮崎市

  • 力学的指標による生細胞表面におけるリガンド‐受容体の相互作用解析
    野坂 和輝, 宮地 佑典, ISHII Jun, OGINO Chiaki, KONDO Akihiko
    化学工学会第42回秋季大会, Sep. 2010, (社)化学工学会, 京都市

  • ガラクトース転写誘導系を利用した酵母リガンド検出系の開発
    漁 慎太郎, 井口 裕介, 福田 展雄, ISHII Jun, KONDO Akihiko
    化学工学会第42回秋季大会, Sep. 2010, (社)化学工学会, 京都市

  • 原子間力顕微鏡を用いた細胞表層におけるリガンド・レセプター間相互作用測定系の構築
    野坂 和輝, 荻野 千秋, 石井 純, 宮地 佑典, 近藤 昭彦
    日本化学会第90春季年会, Mar. 2010, 日本化学会, 東大阪市

  • G蛋白質共役型受容体における二量体化解析のための新規検出システム
    中村 泰之, 石井 純, 近藤 昭彦
    化学工学会第75年会, Mar. 2010, (社)化学工学会, 鹿児島市

  • Split-ubiquitin system for analyzing oligomerization of G protein-coupled receptor
    NAKAMURA Yasuyuki, ISHII Jun, KONDO Akihiko
    APBioChEC'09, Nov. 2009, APBioChEC'09 Committee, 神戸市

  • Investigation of the interaction between GPCR and ligand by AFM equipped with bio-molecule modified cantilever
    NOSAKA K, MIYACHI Y, ISHII Jun, OGINO Chiaki, KONDO Akihiko
    APBioChEC'09, Nov. 2009, English, APBioChEC'09 Committee, 神戸市

  • Functional analysis of mutant human somatostatin receptor using a yeast-based fluorescence reporter assay
    TOGAWA Shota, ISHII Jun, TANAKA Tsutomu, KONDO Akihiko
    APBioChEC'09, Nov. 2009, English, APBioChEC'09 Committee, 神戸市

  • Expression and signaling analyses of human G protein-coupled receptor in yeast
    IGUCHI Yusuke, ISHII Jun, TANAKA Tsutomu, KONDO Akihiko
    APBioChEC'09, Nov. 2009, English, APBioChEC'09 Committee, 神戸市

  • Construction of a novel detection system for protein–protein interactions using yeast G-protein signaling
    FUKUDA Nobuo, ISHII Jun, TANAKA Tsutomu, KONDO Akihiko
    APBioChEC'09, Nov. 2009, English, APBioChEC'09 Committee, 神戸市

  • Bioethaol fermentation from mixed sugar by the recombinant yeast with xyloseisomerase pathway
    TANINO Takanori, HOTTA Atsushi, ITO Tomonori, ISHII Jun, YAMADA Ryousuke, HASUNUMA Tomohisa, OGINO Chiaki, OHMURA Naoto
    APBioChEC'09, Nov. 2009, English, APBioChEC'09 Committee, 神戸市
    Poster presentation

  • 酵母Gタンパク質シグナルを用いたタンパク質間相互作用解析法の開発
    福田 展雄, 石井 純, 田中 勉, 近藤 昭彦
    第61回日本生物工学会大会, Sep. 2009, Japanese, 名古屋市, Domestic conference
    Oral presentation

  • Construction of a novel detection system for protein-protein interactions using yeast G-protein signaling
    Fukuda Nobuo, Ishii Jun, Tanaka Tsutomu, Kondo Akihiko
    The 6th International Forum on Post-Genome Technologies (IFPT'6), Sep. 2009, English, Beijing, China, International conference
    Poster presentation

  • Construction of novel detection system for protein-protein interaction using yeast
    Kondo Akihiko, Fukuda Nobuo, Ishii Jun, Tanaka Tsutomu
    International Workshop HITS 2009, Feb. 2009, English, Tokyo, Japan, International conference
    Poster presentation

  • 酵母G蛋白質共役型受容体蛍光アッセイシステムによるヒトソマトスタチンレセプター細胞外ループドメイン2の変異解析
    Ishii Jun, Matsumura Shizuka, Tsutomu Tanaka, Toshinobu Fujiwara, Tatematsu Kenji, Kuroda Shun'ichi, Fukuda Hideki, Kondo Akihiko
    第31回日本分子生物学会年会・第81回日本生化学会大会合同大会, Dec. 2008, Japanese, 日本分子生物学会年会日本生化学会, 神戸市, Domestic conference
    Poster presentation

  • Mutational analysis for extracellular loop-2 of human somatostatin receptor by yeast-based fluorescence signalling assay
    Ishii Jun, Fujiwara Toshinobu, Kondo Akihiko
    The 14th Symposium of Young Asian Biochemical Engineers' Community (YABEC2008), Nov. 2008, English, Tokyo, Japan, International conference
    Poster presentation

  • Rapid and Efficient Selection of Yeast Displaying a Target Protein Using Thermo-responsive Magnetic Nanoparticles
    Fukuda Nobuo, Ishii Jun, Tanaka Tsutomu, Ohnishi Noriyuki, Fukuda Hideki, Kondo Akihiko
    Bioseparation for Biorecignition and Bionanotechnology Conference, Oct. 2008, English, Ansan, Korea, International conference
    Poster presentation

  • 酵母蛍光レポーターアッセイによるヒトソマトスタチンレセプター変異体の機能解析
    戸川 翔太, 石井 純, 田中 勉, 松村 静香, 立松 健司, 黒田 俊一, 福田 秀樹, 近藤 昭彦
    化学工学会 第40回 秋季大会, Sep. 2008, Japanese, 化学工学会, 宮城県仙台市, Domestic conference
    Poster presentation

  • 酵母でのヒトG蛋白質共役型受容体発現に関する分泌シグナル配列の影響
    井口 裕介, 石井 純, 田中 勉, 立松 健司, 黒田 俊一, 福田 秀樹, 近藤 昭彦
    化学工学会 第40回 秋季大会, Sep. 2008, Japanese, 化学工学会, 宮城県仙台市, Domestic conference
    Poster presentation

  • ペプチド転移酵素を用いた細胞内タンパク質連結技術の開発
    若村 香菜子, 田中 勉, 石井 純, 福田 秀樹, 近藤 昭彦
    化学工学会 第40回 秋季大会, Sep. 2008, Japanese, 化学工学会, 宮城県仙台市, Domestic conference
    Poster presentation

  • Mutational analysis of human somatostatin receptor by yeast-based signalling assay
    Ishii Jun, Kondo Akihiko
    The 3rd International Workshop on Approaches to Single-Cell Analysis, Sep. 2008, English, Zurich, Switzerland, International conference
    Poster presentation

  • ビオチン提示酵母を用いた新規細胞表層提示システムの開発
    増成 伸介, 田中 勉, 石井 純, 福田 秀樹, 近藤 昭彦
    第60回 日本生物工学会2008年度大会, Aug. 2008, Japanese, 日本生物工学会, 宮城県仙台市, Domestic conference
    Oral presentation

  • 酵母ディスプレイ法によるタンパク質の分泌生産/同時回収システム
    Shibasaki Seiji, Kawabata Ai, Ishii Jun, Kadonosono Tetsuya, Maeda Hatsuo, Kondo Akihiko, Ueda Mitsuyoshi
    日本薬学会第128年会, Mar. 2008, Japanese, 日本薬学会, 横浜市, Domestic conference
    Poster presentation

  • 酵母PCAシステムによるタンパク質分子間相互作用の検出
    Sakata Kozue, Sugi Takashi, Ishii Jun, Kondo Akihiko, Ueda Mitsuyoshi, Shibasaki Seiji
    化学工学会第10回化学工学会学生発表会大阪大会(西日本地区), Mar. 2008, Japanese, 化学工学会, 吹田市, Domestic conference
    Oral presentation

  • フローサイトメーター(FCM)を利用した酵母GPCRシグナリング解析技術
    Ishii Jun, Tsutomu Tanaka, Matsumura Shizuka, Tatematsu Kenji, Kuroda Shun'ichi, Ogino Chiaki, Fukuda Hideki, Kondo Akihiko
    化学工学会第73年会, Mar. 2008, Japanese, 化学工学会, 浜松市, Domestic conference
    Oral presentation

  • 酵母GPCRシグナリングにおけるフローサイトメトリー解析技術
    Ishii Jun, Tsutomu Tanaka, Matsumura Shizuka, Tatematsu Kenji, Kuroda Shun'ichi, Ogino Chiaki, Fukuda Hideki, Kondo Akihiko
    第6回最先端バイオテクノロジー若手発表会, Feb. 2008, Japanese, 化学工学会関西支部化学工学会バイオ部会, 西宮市, Domestic conference
    Poster presentation

  • 分子ディスプレイ法によるタンパク質の同時生産/回収システムの構築
    Shibasaki Seiji, Kawabata Ai, Ishii Jun, Kadonosono Tetsuya, Kondo Akihiko, Ueda Mitsuyoshi
    第30回日本分子生物学会年会・第80回日本生化学会大会・合同大会, Dec. 2007, Japanese, 日本分子生物学会年会日本生化学会, 横浜市, Domestic conference
    Oral presentation

  • 酵母を用いたリガンド検出システムの開発
    Moriguchi Miyuki, Ishii Jun, Tatematsu Kenji, Kuroda Shun'ichi, Fukuda Hideki, Kondo Akihiko
    化学工学会第39回秋季大会, Sep. 2007, Japanese, 化学工学会, 札幌市, Domestic conference
    Poster presentation

  • 温度応答性磁性ナノ粒子を用いた迅速かつ高効率なアフィニティ分子選択法の確立
    FUKUDA NOBUO, ISHII JUN, FUKUDA HIDEKI, KONDO AKIHIKO
    第59回 日本生物工学会大会, Sep. 2007, Japanese, 広島大学, Domestic conference
    Poster presentation

  • Fluorescence detection system for human G protein-coupled receptor signaling in yeast
    Ishii Jun, Kondo Akihiko
    The 5th International Forum on Post-Genome Technologies (IFPT'5), Sep. 2007, English, Suzhou, China, International conference
    Poster presentation

  • Fluorescence detection system for heterologous G protein-coupled receptor in yeast
    Ishii Jun, Kondo Akihiko
    The 2nd International Workshop on Approaches to Single-Cell Analysis, Sep. 2007, English, Tokyo, Japan, International conference
    Poster presentation

  • 酵母表層ディスプレイを用いた迅速かつ高効率なタンパク質アフィニティ選択法の確立
    Fukuda Nobuo, Ishii Jun, Shibasaki Seiji, Fukuda Hideki, Kondo Akihiko
    化学工学会第72年会, Mar. 2007, Japanese, 化学工学会, 京都市, Domestic conference
    Oral presentation

  • 蛍光レポーターを用いたヒトGタンパク質共役型受容体に対する高感度な酵母アッセイシステム
    Ishii Jun, Matsumura Shizuka, Tatematsu Kenji, Kuroda Shun'ichi, Fukuda Hideki, Kondo Akihiko
    日本農芸化学会2007年度大会, Mar. 2007, Japanese, 日本農芸化学会, 世田谷区, Domestic conference
    Oral presentation

  • Development of ligand detection system for heterologous G protein-coupled receptor using yeast
    Ishii Jun, Tatematsu Kenji, Kuroda Shun'ichi, Fukuda Hideki, Kondo Akihiko
    The 12th Symposium of Young Asian Biochemical Engineers' Community (YABEC2006), Nov. 2006, English, Kaohsiung, Taiwan, International conference
    Poster presentation

  • 酵母細胞表層ディスプレイを利用した新規タンパク質間相互作用解析システムの開発
    Fukuda Nobuo, Ishii Jun, Shibasaki Seiji, Fukuda Hideki, Kondo Akihiko
    日本生物工学会2006年度大会, Sep. 2006, Japanese, 日本生物工学会, 豊中市, Domestic conference
    Oral presentation

  • 酵母による異種Gタンパク質共役型受容体リガンド検出システムの開発
    Ishii Jun, Matsumura Shizuka, Moriguchi Miyuki, Ishikura Atsushi, Izawa Keiko, Tatematsu Kenji, Kuroda Shun'ichi, Fukuda Hideki, Kondo Akihiko
    日本生物工学会2006年度大会, Sep. 2006, Japanese, 日本生物工学会, 豊中市, Domestic conference
    Oral presentation

  • フェロモン応答性タンパク質を利用した新規レポーターによるリガンド検出システムの開発
    Moriguchi Miyuki, Ishii Jun, Tatematsu Kenji, Kuroda Shun'ichi, Fukuda Hideki, Kondo Akihiko
    化学工学会第38回秋季大会, Sep. 2006, Japanese, 化学工学会, 福岡市, Domestic conference
    Poster presentation

  • Development of a single cell analysis system of agonist for drug discovery
    Kondo Akihiko, Ishii Jun, Matsumura Shizuka, Fukuda Hideki
    The 4th International Forum on Post-Genome Technologies (4’IFPT), Sep. 2006, English, Hangzhou, China, International conference
    Poster presentation

  • Development of a single cell analysis system of agonist for drug discovery
    Kondo Akihiko, Ishii Jun, Matsumura Shizuka, Fukuda Hideki
    The 1st International Workshop on Approaches to Single-Cell Analysis, Jun. 2006, English, Uppsala, Sweden, International conference
    Poster presentation

  • Zドメイン表層提示細胞の機能改変と抗体関連分子の精製への応用
    Yagi Shunsuke, Ishii Jun, Ueda Mitsuyoshi, Kondo Akihiko, Shibasaki Seiji
    日本農芸化学会2006年度大会, Mar. 2006, Japanese, 日本農芸化学会, 京都市, Domestic conference
    Oral presentation

  • EGFP-HIS3融合遺伝子をFUS1遺伝子座に組込んだ酵母細胞におけるシグナル伝達の解析
    Ishii Jun, Matsumura Shizuka, Moriguchi Miyuki, Fukuda Nobuo, Tatematsu Kenji, Kuroda Shun'ichi, Fukuda Hideki, Kondo Akihiko
    第4回最先端バイオテクノロジー公開セミナー, Feb. 2006, Japanese, 化学工学会関西支部化学工学会バイオ部会, 西宮市, Domestic conference
    Poster presentation

  • リガンド・レセプター共提示酵母によるハイスループット創薬システムの開発
    KONDO Akihiko, ISHII Jun
    第6回コンビナトリアル・バイオエンジニアリングシンポジウム, Jan. 2006, Japanese, コンビナトリアル・バイオエンジニアリング研究会, 理化学研究所, Domestic conference
    Invited oral presentation

  • 酵母シグナル伝達経路を利用したリガンドスクリーニングシステムの開発
    Ishii Jun, Matsumura Shizuka, Tatematsu Kenji, Kuroda Shun'ichi, Fukuda Hideki, Kondo Akihiko
    日本生物工学会2005年度大会, Nov. 2005, Japanese, 日本生物工学会, つくば市, Domestic conference
    Oral presentation

  • 酵母シグナル伝達によるリガンド検出システム
    Ishii Jun, Matsumura Shizuka, Tatematsu Kenji, Kuroda Shun'ichi, Fukuda Hideki, Kondo Akihiko
    日本生物工学会2004年度大会, Sep. 2004, Japanese, 日本生物工学会, 名古屋市, Domestic conference
    Oral presentation

Affiliated Academic Society

  • 日本分子生物学会

  • 日本農芸化学会

  • The Society for Biotechnology, Japan

  • 化学工学会

Research Themes

  • 高性能な国産細胞株の構築
    近藤昭彦
    日本医療研究開発機構(AMED), 次世代治療・診断実現のための創薬基盤技術開発事業 / バイオ医薬品の高度製造技術の開発 / 高性能な国産細胞株の構築, 高機能遺伝子デザイン技術研究組合(TRAHED),神戸大学,アステラス製薬,味の素,東京大学,東京工業大学,東北大学, May 2018 - Mar. 2021, Coinvestigator

  • バイオ医薬品の多品種・大量製造に適した微生物による高度生産技術の開発
    石井純
    日本医療研究開発機構(AMED), 次世代治療・診断実現のための創薬基盤技術開発事業 / バイオ医薬品の高度製造技術の開発 / 先端的バイオ製造技術開発, 高機能遺伝子デザイン技術研究組合(TRAHED),神戸大学,Bio-energy, May 2018 - Mar. 2021, Principal investigator

  • 希少アミノ酸エルゴチオネイン高生産スマートセルの開発
    長瀬産業
    新エネルギー・産業技術総合開発機構(NEDO), 植物等の生物を用いた高機能品生産技術の開発 / 微生物による高機能品生産技術開発, 2019 - Feb. 2021, Coinvestigator

  • 高生産性微生物創製に資する情報解析システムの開発
    蓮沼誠久
    新エネルギー・産業技術総合開発機構(NEDO), 植物等の生物を用いた高機能品生産技術の開発, Oct. 2016 - Feb. 2021, Coinvestigator

  • バイオプロダクション次世代農工連携拠点
    福田秀樹, 近藤昭彦
    科学技術振興機構(JST), 先端融合領域イノベーション創出拠点形成プログラム, 神戸大学, 2008 - Mar. 2019, Coinvestigator

  • Matsuda Fumio, ISHII Jun, MORITA Keisuke, NISHINO Shunsuke, NOMURA Yuta, KINOSHITA Shohei
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C), Grant-in-Aid for Scientific Research (C), Osaka University, 01 Apr. 2015 - 31 Mar. 2018
    The central carbon metabolism in budding yeast (Saccharomyces cerevisiae) was engineered to improve the production of butanols from biomass. For the purpose, a precise characterization of yeast metabolism was performed by developing a method to determine concentration of intracellular metabolites as well as to estimate levels of Gibb’s free energy change of metabolic reactionss. The results of the precise analysis suggested that an activation of excess NADH consumption was essential with considering intracellular compartmentalization of yeast cells. Based on the findings, the isobutanol production by yeast was improved by an overexpression of mitochondrial pyruvate carrier proteins.

  • 国際基準に適合した次世代抗体医薬等の製造技術のうち高生産宿主構築の効率化基盤技術の開発に係るもの
    近藤昭彦
    経済産業省(METI) / 日本医療研究開発機構(AMED), 次世代治療・診断実現のための創薬基盤技術開発事業, 高機能遺伝子デザイン技術研究組合(TRAHED),神戸大学,産業技術総合研究所,慶應義塾大学,カネカ,アステラス製薬,プレシジョン・システム・サイエンス,インシリコバイオロジー,東京工業大学,東北大学,東京大学,弘前大学,金澤工業大学,大阪府立大学, Oct. 2014 - Mar. 2018, Coinvestigator

  • 光駆動ATP再生系によるVmax細胞の創製
    原清敬
    科学技術振興機構(JST), 未来社会創造事業(MIRAI), Nov. 2017, Coinvestigator
    Competitive research funding

  • 革新的バイオマテリアル実現のための高機能化ゲノムデザイン技術開発
    近藤昭彦
    経済産業省(METI), 経済産業省委託事業, 高機能遺伝子デザイン技術研究組合(TRAHED),神戸大学,産業技術総合研究所,慶應義塾大学,プレシジョン・システム・サイエンス,Spiber,小島プレス工業,味の素,三菱化学,カネカ,神戸天然物化学,アステラス製薬,インシリコバイオロジー,クミアイ化学工業,次世代天然物化学技術研究組合,東北大学,北海道大学,京都大学,国立遺伝学研究所,千葉大学,東京工業大学,石川県立大学,鳥取大学,理化学研究所,バイオインダストリー協会, Oct. 2012 - Mar. 2017, Coinvestigator

  • インドネシアにおける統合バイオリファイナリーシステムの開発
    荻野千秋
    国際協力機構(JICA) / 科学技術振興機構(JST), 地球規模課題対応国際科学技術協力(SATREPS), 神戸大学,インドネシア科学院(LIPI), Apr. 2012 - Mar. 2017, Coinvestigator

  • 【NEDO:三菱ケミカル】共同研究員受入
    近藤 昭彦
    三菱ケミカル株式会社, NEDO委託プロジェクト「植物等の生物を用いた高機能品生産技術の開発」, 2017
    Competitive research funding

  • Ishii Jun
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for Young Scientists (B), Grant-in-Aid for Young Scientists (B), Kobe University, Apr. 2014 - Mar. 2016, Principal investigator
    As the switching genome editing technology that can respond to G-protein signaling, we established the method permitting the pop-out of a gene and the alternative expression of another gene based on the Cre/loxP recombination technique. Herewith, it has enabled to induce concurrently “ON to OFF” and “OFF to ON” of two genes in concert with the detection of G-protein signaling. Using this technology, we developed the system for switching two gene expressions in response to the signal for dimer formation of G-protein coupled receptors (GPCRs); thereby, successfully detecting the homodimer formations of yeast endogenous Ste2 receptor and human serotonin receptor (HTR1A), and the heterodimer formation of human adrenergic receptor (ADRB2).
    Competitive research funding

  • 新規代謝デザインにもとづく次世代バイオ燃料(イソブタノール)生産酵母の開発
    松田史生
    新エネルギー・産業技術総合開発機構(NEDO), 先導的産業技術創出事業費助成金, 大阪大学,神戸大学, Oct. 2011 - Sep. 2015, Coinvestigator

  • MATSUDA Fumio, ISHII Jun, MORITA Keisuke, NOMURA Yuta
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for Young Scientists (B), Grant-in-Aid for Young Scientists (B), Osaka University, 01 Apr. 2013 - 31 Mar. 2015
    The central carbon metabolism in baker's yeast, Saccharomyces cerevisiae, was expanded by introducing metabolic pathways of other microorganisms to improve a capability of the yeast metabolism for more efficient bioproduction of useful chemicals. The metabolically engineered S. cerevisiae strains expressing bacterial Entner-Doudrouf and phosphoenolpyruvate carboxylase pathways were constructed whose functions were confirmed by the labeling experiment and isobutanol fermentaion test.

  • 酵母細胞表層工学によるエタノール・マンノオリゴ糖生産の統合バイオプロセス開発
    荻野千秋
    日本学術振興会(JSPS), 二国間交流事業共同研究, 神戸大学,インドネシア科学院(LIPI), 2011 - 2013, Coinvestigator

  • 7回膜受容体シグナル伝達機構解析のための新規バイオセンサの開発
    石井純
    公益財団法人 内藤記念科学振興財団, 第42回内藤記念科学奨励金・研究助成, 神戸大学, Oct. 2010 - Sep. 2012, Principal investigator

  • 原子間力顕微鏡を利用した力学的生物界面のナノスケール現象解析
    荻野千秋
    科学技術振興機構(JST), 研究シーズ探索プログラム(融合分野), 神戸大学, 2009 - 2010, Coinvestigator

Industrial Property Rights

  • 新規細胞及びそれを用いた目的タンパク質の製造方法
    伊藤洋一郎, 石井純, 中村泰之, 近藤昭彦, 西輝之
    特願2018-058953, 26 Mar. 2018, 国立大学法人神戸大学, 株式会社カネカ, 特許第7349684号, 14 Sep. 2023
    Patent right

  • 新規宿主細胞及びそれを用いた目的タンパク質の製造方法
    伊藤 洋一郎, 石井 純, 中村 泰之, 近藤 昭彦, 西 輝之
    特願2016-172841, 05 Sep. 2016, 国立大学法人神戸大学, 特開2021-101733, 15 Jul. 2021, 特許第7239205号, 06 Mar. 2023
    Patent right

  • キメラプラスミドライブラリーの構築方法
    柘植謙爾, 石井純, 近藤昭彦
    特願2019-069798, 01 Apr. 2019, 国立大学法人神戸大学, 特許第7101431号, 07 Jul. 2022
    Patent right

  • 遺伝子スイッチ
    冨永 将大, 石井 純, 能崎 健太, 近藤 昭彦
    特願2020-202234, 04 Dec. 2020, 国立大学法人神戸大学, 特開2022-089660, 16 Jun. 2022
    Patent right

  • 二重特異性抗体
    中村泰之, 藍川晋平, 伊藤洋一郎, 西村勇哉, 足達哲也, 石井純, 近藤昭彦, 西輝之, 梅津光央, 中澤光, 杉山在生人
    特願2017-237252, 11 Dec. 2017, 国立大学法人神戸大学, 株式会社カネカ, 国立大学法人東北大学, 特開2019-104699, 27 Jun. 2019, 特許第7072792号, 13 May 2022, 23 May 2022
    Patent right

  • 遺伝子スイッチを選抜する方法
    冨永将大, 石井純, 能崎健太, 近藤昭彦
    特願2020-202233, 04 Dec. 2020, 国立大学法人神戸大学
    Patent right

  • 高発現高制御性遺伝子スイッチ
    冨永将大, 石井純, 伊藤洋一郎, 能崎健太, 近藤昭彦
    特願2020-202239, 04 Dec. 2020, 国立大学法人神戸大学
    Patent right

  • 外来性のオプシンおよび当該ロドプシンを発現した細胞
    原清敬, 原陽子, 戸谷吉博, 松田史生, 田中涼, 戸谷さや香, 佐野海瑚人, 石井純, 柘植謙爾, 平山英伸, 木苗貴秀, 武田泉穂, 菊川寛史, 酒井香奈江
    特願2020-140730, 24 Aug. 2020, 静岡県公立大学法人静岡県立大学, 国立大学法人大阪大学, 国立大学法人神戸大学, 株式会社けんだいパートナーズ
    Patent right

  • 内在性遺伝子の発現が増強された遺伝子組換え細胞の製造方法
    伊藤洋一郎, 石井純, 近藤昭彦
    特願2020-050192, 19 Mar. 2020, 国立大学法人神戸大学
    Patent right

  • ニコチンアミド誘導体を製造するための組換え微生物及び方法、並びにそれに用いられるベクター
    庄司信一郎, 石井純, 近藤昭彦, 狩野理延, 中島良太, 渡邉秀和
    特願2018-236634, 18 Dec. 2018, 国立大学法人神戸大学, 帝人株式会社
    Patent right

  • 新規宿主細胞及びそれを用いた目的タンパク質の製造方法
    伊藤洋一郎, 石井純, 中村泰之, 近藤昭彦, 西輝之
    特願2016-172841, 05 Sep. 2016, 国立大学法人神戸大学, 株式会社カネカ, 特開2018-38286, 15 Mar. 2018
    Patent right

  • イソブタノール生産酵母
    松田史生, 近藤昭彦, 石井純
    特願2013-94568, 26 Apr. 2013, 国立大学法人神戸大学, 特開2014-212762, 17 Nov. 2014
    Patent right

  • 高親和性のタンパク質間相互作用検出・スクリーニング方法
    福田 展雄, ISHII JUN, TANAKA TSUTOMU, KONDO AKIHIKO
    特願2010-017509, 28 Jan. 2010, 国立大学法人神戸大学, 特許5574479, 11 Jul. 2014
    Patent right

  • レセプター結合性物質のスクリーニング方法
    近藤昭彦, 黒田俊一, 植田充美, 石井純, 福田秀樹, 立松健司
    特願2003-417482, 16 Dec. 2003, バイオ・エナジー株式会社, 特許第5224491号, 22 Mar. 2013
    Patent right

  • イソプロピルアルコール生産酵母及びイソプロピルアルコール生産方法
    松本佳子, 竹林のぞみ, 高橋均, 和田光史, 近藤昭彦, 石井純, 近藤貴志
    特願2010-226668, 06 Oct. 2010, 三井化学株式会社, 国立大学法人神戸大学, 特開2011-97929, 19 May 2011
    Patent right