研究者紹介システム

秀瀨 涼太
ヒデセ リョウタ
大学院科学技術イノベーション研究科 科学技術イノベーション専攻
准教授
農芸化学関係
Last Updated :2021/07/25

研究者情報

所属

  • 【主配置】

    大学院科学技術イノベーション研究科 科学技術イノベーション専攻
  • 【配置】

    先端バイオ工学研究センター

学位

  • 博士(農学), 京都大学

ジャンル

  • 科学・技術 / バイオテクノロジー

研究活動

研究分野

  • ライフサイエンス / 分子生物学
  • ライフサイエンス / 応用微生物学

受賞

  • 2019年 日本生物工学会, 生物工学論文賞, Leucine responsive regulatory protein is involved in methionine metabolism and polyamine homeostasis in acetic acid bacterium Komagataeibacter europaeus

    Yuri Ishii, Naoki Akasaka, Hisao Sakoda, Ryota Hidese, Shinsuke Fujiwara

  • 2018年 日本生物工学会, 2018年度 第26回 生物工学論文賞

    秀瀨 涼太

  • 2018年 長瀬科学技術振興財団, 平成30年度 長瀬研究振興賞

    秀瀨 涼太

  • 2018年 極限環境生物学会, 研究奨励賞

    秀瀨 涼太

論文

  • Yuichi Kato, Tomoki Oyama, Kentaro Inokuma, Christopher J. Vavricka, Mami Matsuda, Ryota Hidese, Katsuya Satoh, Yutaka Oono, Jo-Shu Chang, Tomohisa Hasunuma, Akihiko Kondo

    AbstractLight/dark cycling is an inherent condition of outdoor microalgae cultivation, but is often unfavorable for lipid accumulation. This study aims to identify promising targets for metabolic engineering of improved lipid accumulation under outdoor conditions. Consequently, the lipid-rich mutant Chlamydomonas sp. KOR1 was developed through light/dark-conditioned screening. During dark periods with depressed CO2 fixation, KOR1 shows rapid carbohydrate degradation together with increased lipid and carotenoid contents. KOR1 was subsequently characterized with extensive mutation of the ISA1 gene encoding a starch debranching enzyme (DBE). Dynamic time-course profiling and metabolomics reveal dramatic changes in KOR1 metabolism throughout light/dark cycles. During light periods, increased flux from CO2 through glycolytic intermediates is directly observed to accompany enhanced formation of small starch-like particles, which are then efficiently repartitioned in the next dark cycle. This study demonstrates that disruption of DBE can improve biofuel production under light/dark conditions, through accelerated carbohydrate repartitioning into lipid and carotenoid.

    Springer Science and Business Media LLC, 2021年12月, Communications Biology, 4 (1)

    研究論文(学術雑誌)

  • Ryota Hidese, Mami Matsuda, Takashi Osanai, Tomohisa Hasunuma, Akihiko Kondo

    d-Lactate is one of the most valuable compounds for manufacturing biobased polymers. Here, we have investigated the significance of endogenous malate dehydrogenase (decarboxylating) (malic enzyme, ME), which catalyzes the oxidative decarboxylation of malate to pyruvate, in d-lactate biosynthesis in the cyanobacterium Synechocystis sp. PCC6803. d-Lactate levels were increased by 2-fold in ME-overexpressing strains, while levels in ME-deficient strains were almost equivalent to those in the host strain. Dynamic metabolomics revealed that overexpression of ME led to increased turnover rates in malate and pyruvate metabolism; in contrast, deletion of ME resulted in increased pool sizes of glycolytic intermediates, probably due to sequential feedback inhibition, initially triggered by malate accumulation. Finally, both the loss of the acetate kinase gene and overexpression of endogenous d-lactate dehydrogenase, concurrent with ME overexpression, resulted in the highest production of d-lactate (26.6 g/L) with an initial cell concentration of 75 g-DCW/L after 72 h fermentation.

    2020年02月21日, ACS synthetic biology, 9 (2), 260 - 268, 英語, 国際誌

    [査読有り]

  • Wakao Fukuda, Yuka Yamori, Masafumi Hamakawa, Mamoru Osaki, Moeko Fukuda, Ryota Hidese, Yu Kanesaki, Akiko Okamoto-Kainuma, Satoru Kato, Shinsuke Fujiwara

    Branched-chain polyamine (BCPA) synthase (BpsA), encoded by the bpsA gene, is responsible for the biosynthesis of BCPA in the hyperthermophilic archaeon Thermococcus kodakarensis, which produces N4-bis(aminopropyl)spermidine and spermidine. Here, next-generation DNA sequencing and liquid chromatography-mass spectrometry (LC-MS) were used to perform transcriptomic and proteomic analyses of a T. kodakarensis strain (DBP1) lacking bpsA. Subsequently, the contributions of BCPA to gene transcription (or transcript stabilization) and translation (or protein stabilization) were analyzed. Compared with those in the wild-type strain (KU216) cultivated at 90 °C, the transcript levels of 424 and 21 genes were up- and downregulated in the DBP1 strain, respectively. The expression levels of 12 frequently-used tRNAs were lower in DBP1 cells than KU216 cells, suggesting that BCPA affects translation efficiency in T. kodakarensis. LC-MS analyses of cells grown at 90 °C detected 50 proteins in KU216 cells only, 109 proteins in DBP1 cells only, and 499 proteins in both strains. Notably, the transcript levels of some genes did not correlate with those of the proteins. RNA-seq and RT-qPCR analyses of ten proteins that were detected in KU216 cells only, including three flagellin-related proteins (FlaB2-4) and cytosolic NiFe-hydrogenase subunit alpha (HyhL), revealed that the corresponding transcripts were expressed at higher levels in DBP1 cells than KU216 cells. Electron microscopy analyses showed that flagella formation was disrupted in DBP1 cells at 90 °C, and western blotting confirmed that HyhL expression was eliminated in the DBP1 strain. These results suggest that BCPA plays a regulatory role in gene expression in T. kodakarensis.

    Springer Science and Business Media LLC, 2020年02月, Amino acids, 52 (2), 287 - 299, 英語, 国際誌

    [査読有り]

    研究論文(学術雑誌)

  • The C-terminal flexible region of branched-chain polyamine synthase facilitates substrate specificity and catalysis

    Ryota Hidese, Masataka Toyoda, Ken-ichi Yoshino, Wakao Fukuda, Gita Adhirani Wihardja, Seigo Kimura, Junso Fujita, Masaru Niitsu, Tairo Oshima, Tadayuki Imanaka, Eiichi Mizohata, Shinsuke Fujiwara

    2019年10月, The FEBS Journal, 286 (19), 3926 - 3940, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Branched-chain polyamine stabilizes RNA polymerase at elevated temperatures in hyperthermophiles

    YAMORI Yuka, HAMAKAWA Masafumi, HIDESE Ryota, FUKUDA Moeko, ATOMI Haruyuki, FUKUDA Wakao, FUJIWARA Shinsuke

    2019年05月, Amino Acids, 1 - 11, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Increased flux in acetyl-CoA synthetic pathway and TCA cycle of Kluyveromyces marxianus under respiratory conditions

    SAKIHAMA Yuri, HIDESE Ryota, HASUNUMA Tomohisa, KONDO Akihiko

    2019年03月, Scientific Reports, 9 (1), 5319, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Accurate fidelity analysis of the reverse transcriptase by a modified next-generation sequencing.

    OKANO Hiroyuki, BABA Misato, HIDESE Ryota, IIDA Kei, LI Tongyang, KOJIMA Kenji, TAKITA Teisuke, YANAGIHARA Itaru, FUJIWARA Shinsuke, YASUKAWA Kiyoshi

    2018年08月, Enzyme and Microbial Technology, 115, 81 - 85, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Agmatine Production by Aspergillus oryzae Is Elevated by Low pH during Solid-State Cultivation

    AKASAKA Naoki, KATO Saori, KATO Saya, HIDESE Ryota, WAGU Yutaka, SAKODA Hisao, FUJIWARA Shinsuke

    2018年07月, Applied and Environmental Microbiology, 17 (84), 15, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Hiroyuki Okano, Misato Baba, Katsuhiro Kawato, Ryota Hidese, Itaru Yanagihara, Kenji Kojima, Teisuke Takita, Shinsuke Fujiwara, Kiyoshi Yasukawa

    One-step RT-PCR has not been widely used even though some thermostable DNA polymerases with reverse transcriptase (RT) activity were developed from bacterial and archaeal polymerases, which is owing to low cDNA synthesis activity from RNA. In the present study, we developed highly-sensitive one-step RT-PCR using the single variant of family A DNA polymerase with RT activity, K4polL329A (L329A), from the hyperthermophilic bacterium Thermotoga petrophila K4 or the 16-tuple variant of family B DNA polymerase with RT activity, RTX, from the hyperthermophilic archaeon Thermococcus kodakarensis. Optimization of reaction condition revealed that the activities for cDNA synthesis and PCR of K4polL329A and RTX were highly affected by the concentrations of MgCl2 and Mn(OCOCH3)2 as well as those of K4polL329A or RTX. Under the optimized condition, 300 copies/μl of target RNA in 10 μl reaction volumes were successfully detected by the one-step RT-PCR with K4polL329A or RTX, which was almost equally sensitive enough compared with the current RT-PCR condition using retroviral RT and thermostable DNA polymerase. Considering that K4polL329A and RTX are stable even at 90-100°C, our results suggest that the one-step RT-PCR with K4polL329A or RTX is more advantageous than the current one.

    2018年03月, Journal of bioscience and bioengineering, 125 (3), 275 - 281, 英語, 国内誌

    [査読有り]

    研究論文(学術雑誌)

  • Ryota Hidese, Wakao Fukuda, Masaru Niitsu, Shinsuke Fujiwara

    Thermophiles are organisms that grow optimally at temperatures higher than 55 °C. They contain two types of unusual longer/branched-chain polyamines in addition to common polyamines such as spermidine and putrescine. These unusual polyamines contribute to the survival of hyperthermophiles at high temperatures. Recently, the novel aminopropyltransferase BpsA was found to be responsible for the biosynthesis of branched-chain polyamines in the hyperthermophilic archaeon Thermococcus kodakarensis, which contains N4-bis(aminopropyl)spermidine as the major polyamine. This compound is synthesized by the sequential addition of decarboxylated S-adenosylmethionine (dcSAM) aminopropyl groups to spermidine via the bifunctional catalytic action of BpsA. In this chapter, methods for the extraction and identification of branched-chain polyamines are presented, along with methods for the production and characterization of recombinant T. kodakarensis BpsA as a model aminopropyltransferase.

    Humana Press Inc., 2018年, Methods in Molecular Biology, 1694, 81 - 94, 英語

    [査読有り]

    論文集(書籍)内論文

  • Thermostable DNA helicase improves the sensitivity of digital PCR

    HIDESE Ryota, KAWATO Katsuhiro, NAKURA Yukiko, FUJIWARA Ayako, YASUKAWA Kiyoshi, YANAGIHARA Itaru, FUJIWARA Shinsuke

    2018年01月, Biochemical and Biophysical Research Communications, 495 (3), 2189 - 2194, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Yuri Ishii, Naoki Akasaka, Hisao Sakoda, Ryota Hidese, Shinsuke Fujiwara

    The leucine responsive regulatory protein (Lrp) is a global transcription factor that regulates the expression of genes involved in amino acid metabolism. To identify metabolic pathways and related genes under the control of Lrp in the acetic acid bacterium Komagataeibacter europaeus, the Kelrp null mutant (KGMA7110), which requires supplementation of all 20 amino acids for normal growth, was cultivated in minimal media containing or lacking particular amino acids. The results confirmed that KGMA7110 was auxotrophic for methionine and its catabolites S-adenosylmethionine (SAM) and spermidine (SPD). Quantitative reverse-transcription PCR analysis revealed lower metK (SAM synthetase) and mdtI (SPD efflux pump) expression in KGMA7110 than in wild-type KGMA0119. By contrast, these genes were significantly up-regulated in the Kelrp mutant lacking the putative C-terminal ligand-sensing domain (KGMA7203), indicating abnormal regulation of target genes by the KeLrp variant in KGMA7203. KGMA7110 (0.69 ± 0.27 μM) and KGMA7203 (4.90 ± 0.61 μM) excreted lower and higher quantities of SPD, respectively, than KGMA0119 (2.28 ± 0.26 μM). This was attributed to imbalanced carbon flow caused by Kelrp disruption that respectively attenuated and stimulated metK and mdtI expression. These findings indicate that KeLrp plays a key role in SAM biosynthesis and intracellular polyamine homeostasis in K. europaeus.

    Elsevier B.V., 2018年01月01日, Journal of Bioscience and Bioengineering, 125 (1), 67 - 75, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Ryota Hidese, Ka Man Tse, Seigo Kimura, Eiichi Mizohata, Junso Fujita, Yuhei Horai, Naoki Umezawa, Tsunehiko Higuchi, Masaru Niitsu, Tairo Oshima, Tadayuki Imanaka, Tsuyoshi Inoue, Shinsuke Fujiwara

    Branched-chain polyamines are found exclusively in thermophilic bacteria and Euryarchaeota and play essential roles in survival at high temperatures. In the present study, kinetic analyses of a branched-chain polyamine synthase from the hyperthermophilic archaeon Thermococcus kodakarensis (Tk-BpsA) were conducted, showing that N-4-bis(aminopropyl)spermidine was produced by sequential additions of decarboxylated S-adenosylmethionine (dcSAM) aminopropyl groups to spermidine, through bifunctional catalytic action. Tk-BpsA catalyzed the aminopropylation of the linear-chain polyamines spermidine, spermine, norspermidine, and the tertiary-branched polyamines N-4-aminopropylspermidine and N-4-aminopropylnorspermidine, but not of short-chain diamines, putrescine, and cadaverine, suggesting that Tk-BpsA does not catalyze the aminopropylation of primary amino groups of diamines. X-ray structural analyses of Tk-BpsA in the presence or absence of the substrates spermidine and dcSAM revealed that a large, negatively charged cavity is responsible for the binding of branched-chain substrates. The binding is different from that in the active site of linear polyamine spermidine/spermine synthases, and loop-closures occur upon the binding of spermidine. Based on structural analyses, further kinetic studies were carried out for various mutants, revealing that Asp159, positioned between the reactive secondary amino group of the substrate polyamine and a sulfur atom of the product 5'-methylthioadenosine and in a Gly-Asp-Asp-Asp motif, functions as a catalytic center, with reactions proceeding via a ping-pong mechanism. Our study provides a novel aminopropyltransfer reaction mechanism, distinct from the S(N)2 displacement mechanism found in other known linear spermidine/spermine synthases. DatabaseAtomic coordinates and structure factors have been deposited in the Protein Data Bank with PDB codes for apo-Tk-BpsA, 5XNH for the binary complex, and 5XNC for the ternary complex.

    WILEY, 2017年11月, FEBS JOURNAL, 284 (21), 3684 - 3701, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Kiyoshi Yasukawa, Kei Iida, Hiroyuki Okano, Ryota Hidese, Misato Baba, Itaru Yanagihara, Kenji Kojima, Teisuke Takita, Shinsuke Fujiwara

    In this study, we devised a simple and rapid method to analyze fidelity of reverse transcriptase (RT) using next-generation sequencing (NGS). The method comprises a cDNA synthesis reaction from standard RNA with a primer containing a tag of 14 randomized bases and the RT to be tested, PCR using high-fidelity DNA polymerase, and NGS. By comparing the sequence of each read with the reference sequence, mutations were identified. The mutation can be identified to be due to an error introduced by either cDNA synthesis, PCR, or NGS based on whether the sequence reads with the same tag contain the same mutation or not. The error rates in cDNA synthesis with Moloney murine leukemia virus (MMLV) RT thermostable variant MM4 or the recently developed 16-tuple variant of family B DNA polymerase with RT activity, RTX, from Thermococcus kodakarensis, were 0.75-1.0 x 10(-4) errors/base, while that in the reaction with the wild-type human immunodeficiency virus type I (HIV-1) RT was 2.6 x 10(-4) errors/base. Overall, our method could precisely evaluate the fidelity of various RTs with different reaction conditions in a high-throughput manner without the use of expensive optics and troublesome adaptor ligation. (C) 2017 Elsevier Inc. All rights reserved.

    ACADEMIC PRESS INC ELSEVIER SCIENCE, 2017年10月, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 492 (2), 147 - 153, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Ryota Hidese, Keita Yamashita, Kohei Kawazuma, Tamotsu Kanai, Haruyuki Atomi, Tadayuki Imanaka, Shinsuke Fujiwara

    The redox-responsive regulator SurR in the hyperthermophilic archaea Pyrococcus furiosus and Thermococcus kodakarensis binds to the SurR-binding consensus sequence (SBS) by responding to the presence of elemental sulfur. Here we constructed a surR gene disruption strain (DTS) in T. kodakarensis, and identified the genes that were under SurR control by comparing the transcriptomes of DTS and parent strains. Among these genes, transcript levels of ferredoxin:NADP(+) oxidoreductases 1 and 2 (FNOR1 and FNOR2) genes displayed opposite responses to surR deletion, indicating that SurR repressed FNOR1 transcription while enhancing FNOR2 transcription. Each promoter region contains an SBS upstream (uSBS) and downstream (dSBS) of TATA. In addition to in vitro binding assays, we examined the roles of each SBS in vivo. In FNOR1, mutations in either one of the SBSs resulted in a complete loss of repression, indicating that the presence of both SBSs was essential for repression. In FNOR2, uSBS indeed functioned to enhance gene expression, whereas dSBS functioned in gene repression. SurR bound to uSBS2 of FNOR2 more efficiently than to dSBS2 in vitro, which may explain why SurR overall enhances FNOR2 transcription. Further analyses indicated the importance in the distance between uSBS and TATA for transcriptional activation in FNOR2.

    SPRINGER JAPAN KK, 2017年09月, EXTREMOPHILES, 21 (5), 903 - 917, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Le Gao, Ryota Hidese, Shinsuke Fujiwara

    Molecular chaperonin CpkB from Thermococcus kodakarensis possesses a unique negatively charged carboxy-terminal region that functions in target protein recognition. In the present study, green fluorescent protein (GFP), 4-oxalocrotonate tautomerase (4OTA) and glutamine:fructose-6-phosphate amidotransferase (GFAT) were fused with a positively charged tag, selected using docking simulation in silico, to enhance their electrostatic interactions with CpkB. Target proteins were heated at 75 degrees C in the presence or absence of CpkB, and the remaining enzymatic activity was measured. The half-life (t(1/2)) of the positively charged tagged targets was significantly longer than that of their tagless counterparts. Escherichia coli cell extracts containing heterologously expressed targets (GFP, 4OTA and GFAT and their tagged variants) were incubated at 75 degrees C in the presence or absence of CpkB, and the proportion remaining in the soluble fraction was evaluated by SDS-PAGE. Only positively charged tagged targets remained predominantly in the soluble fraction in the presence of CpkB but not in the absence of CpkB. When tagless or negatively charged tagged targets were employed, the targets were barely detected in the soluble fraction, suggesting that CpkB protected positively charged tagged proteins more efficiently than tagless targets. Attachment of a positively charged tag may be a generally applicable method for enhancing target recognition by chaperonins carrying negatively charged carboxy-terminal regions, such as the archaeal chaperonin CpkB. (C) 2017, The Society for Biotechnology, Japan. All rights reserved.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2017年09月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 124 (3), 283 - 288, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Hiroyuki Okano, Misato Baba, Tomomi Yamasaki, Ryota Hidese, Shinsuke Fujiwara, Itaru Yanagihara, Takeshi Ujiiye, Tsukasa Hayashi, Kenji Kojima, Teisuke Takita, Kiyoshi Yasukawa

    ACADEMIC PRESS INC ELSEVIER SCIENCE, 2017年05月, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 487 (1), 128 - 133, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Hiroyuki Okano, Yuta Katano, Misato Baba, Ayako Fujiwara, Ryota Hidese, Shinsuke Fujiwara, Itaru Yanagihara, Tsukasa Hayashi, Kenji Kojima, Teisuke Takita, Kiyoshi Yasukawa

    Detection of mRNA is a valuable method for monitoring the specific gene expression. In this study, we devised a novel cDNA synthesis method using three enzymes, the genetically engineered thermostable variant of reverse transcriptase (RT), MM4 (E28612/E302K/L435R/D524A) from Moloney murine leukemia virus (MMLV), the genetically engineered variant of family A DNA polymerase with RT activity, K4pol(L329A) from thermophilic Thermotoga petrophila K4, and the DNA/RNA helicase Tk-EshA from a hyperthermophilic archaeon Thermococcus kodakarensis. By optimizing assay conditions for three enzymes using Taguchi's method, 100 to 1000-fold higher sensitivity was achieved for cDNA synthesis than conventional assay condition using only RT. Our results suggest that DNA polymerase with RT activity and DNA/RNA helicase are useful to increase the sensitivity of cDNA synthesis. (C) 2016 Elsevier Inc. All rights reserved.

    ELSEVIER SCIENCE INC, 2017年01月, ENZYME AND MICROBIAL TECHNOLOGY, 96, 111 - 120, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Ryota Hidese, Ki-Hwan Im, Masaki Kobayashi, Masaru Niitsu, Takemitsu Furuchi, Shinsuke Fujiwara

    Long/branched-chain polyamines are unique polycations found in thermophiles. The hyperthermophilic archaeon Thermococcus kodakarensis contains spermidine and a branched-chain polyamine, N-4-bis(aminopropyl)spermidine, as major polyamines. The metabolic pathways associated with branched-chain polyamines remain unknown. Here, we used gas chromatography and liquid chromatography-tandem mass spectrometry analyses to identify a new acetylated polyamine, N-4-bis(aminopropyl)-N-1-acetylspermidine, from T. kodakarensis; this polyamine was not found in other micro-organisms. The amounts of branched-chain polyamine and its acetylated form increased with temperature, indicating that branched-chain polyamines are important for growth at higher temperatures. The amount of quaternary acetylated polyamine produced was associated with the amount of N-4-bis(aminopropyl)spermidine in the cell. The ratio of acetylated to non-acetylated forms was higher in the stationary phase than in the logarithmic growth phase under high-temperature stress condition.

    TAYLOR & FRANCIS LTD, 2017年, BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY, 81 (9), 1845 - 1849, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Application of a Euryarchaeota-Specific Helicase from Thermococcus kodakarensis for Noise Reduction in PCR

    Ayako Fujiwara, Katsuhiro Kawato, Saori Kato, Kiyoshi Yasukawa, Ryota Hidese, Shinsuke Fujiwara

    DNA/RNA helicases, which are enzymes for eliminating hydrogen bonds between bases of DNA/DNA, DNA/RNA, and RNA/ RNA using the energy of ATP hydrolysis, contribute to various biological activities. In the present study, the Euryarchaeota-specific helicase EshA (TK0566) from the hyperthermophilic archaeon Thermococcus kodakarensis (Tk-EshA) was obtained as a recombinant form, and its enzymatic properties were examined. Tk-EshA exhibited maximal ATPase activity in the presence of RNA at 80 degrees C. Unwinding activity was evaluated with various double-stranded DNAs (forked, 5' overhung, 3' overhung, and blunt end) at 50 degrees C. Tk-EshA unwound forked and 3' overhung DNAs. These activities were expected to unwind the structured template and to peel off misannealed primers when Tk-EshA was added to a PCR mixture. To examine the effect of Tk-EshA on PCR, various target DNAs were selected, and DNA synthesis was investigated. When 16S rRNA genes were used as a template, several misamplified products (noise DNAs) were detected in the absence of Tk-EshA. In contrast, noise DNAs were eliminated in the presence of Tk-EshA. Noise reduction by Tk-EshA was also confirmed when Taq DNA polymerase (a family A DNA polymerase, PolI type) and KOD DNA polymerase (a family B DNA polymerase, alpha type) were used for PCR. Misamplified bands were also eliminated during toxA gene amplification from Pseudomonas aeruginosa DNA, which possesses a high GC content (69%). Tk-EshA addition was more effective than increasing the annealing temperature to reduce misamplified DNAs during toxA amplification. Tk-EshA is a useful tool to reduce noise DNAs for accurate PCR. IMPORTANCE PCR is a technique that is useful for genetic diagnosis, genetic engineering, and detection of pathogenic microorganisms. However, troubles with nonspecific DNA amplification often occur from primer misannealing. In order to achieve a specific DNA amplification by eliminating noise DNAs derived from primer misannealing, a thermostable Euryarchaeota-specific helicase (Tk-EshA) was included in the PCR mixture. The addition of Tk-EshA has reduced noise DNAs in PCR.

    AMER SOC MICROBIOLOGY, 2016年05月, APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 82 (10), 3022 - 3031, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Hidese Ryota, Fukuda Wakao, Fujiwara Shinsuke, Kanai Tamotsu, Atomi Haruyuki, Imanaka Tadayuki

    2016年, Gene Expression Omnibus

    [査読有り]

  • CuI and H2O2 Inactivate and FeII Inhibits [Fe]-Hydrogenase at very low Concentrations

    HIDESE Ryota, ATAKA Kenichi, BILL Eckhard, SHIMA Seigo

    2015年07月, ChemBioChem, 16 (13), 1861 - 1865, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Naoki Akasaka, Wiwik Astuti, Yuri Ishii, Ryota Hidese, Hisao Sakoda, Shinsuke Fujiwara

    Plasmids pGE1 (2.5 kb), pGE2 (7.2 kb), and pGE3 (5.5 kb) were isolated from Gluconacetobacter europaeus KGMA0119, and sequence analyses revealed they harbored 3, 8, and 4 genes, respectively. Plasmid copy numbers (PCNs) were determined by real-time quantitative PCR at different stages of bacterial growth. When KGMA0119 was cultured in medium containing 0.4% ethanol and 0.5% acetic acid, PCN of pGE1 increased from 7 copies/genome in the logarithmic phase to a maximum of 12 copies/genome at the beginning of the stationary phase, before decreasing to 4 copies/genome in the late stationary phase. PCNs for pGE2 and pGE3 were maintained at 1-3 copies/genome during all phases of growth. Under a higher concentration of ethanol (3.2%) the PCN for pGE1 was slightly lower in all the growth stages, and those of pGE2 and pGE3 were unchanged. In the presence of 1.0% acetic acid, PCNs were higher for pGE1 (10 copies/genome) and pGE3 (6 copies/genome) during the logarithmic phase. Numbers for pGE2 did not change, indicating that pGE1 and pGE3 increase their PCNs in response to acetic acid. Plasmids pBE2 and pBE3 were constructed by ligating linearized pGE2 and pGE3 into pBR322. Both plasmids were replicable in Escherichia coli, Acetobacter pasteurianus and G. europaeus, highlighting their suitability as vectors for acetic acid bacteria. (C) 2014, The Society for Biotechnology, Japan. All rights reserved.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2015年06月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 119 (6), 661 - 668, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Wakao Fukuda, Ryota Hidese, Shinsuke Fujiwara

    Long-chain and/or branched polyamines such as N 4 - aminopropylspermidine [3(3)4] (abbreviation for the number of methylene CH 2 chain units between NH 2, NH, N, or N +), N 4 -bis(aminopropyl)spermidine [3(3) (3)4], and tetrakis(3-aminopropyl)ammonium [3(3)(3)3] are polycations of biotic origin that are only found in thermophiles. The thermophilic bacterium Thermus thermophilus and the hyperthermophilic archaeon Thermococcus kodakarensis synthesize a polyamine, spermidine, via conversion of arginine to agmatine (a step catalyzed by arginine decarboxylase), aminopropylation of agmatine to N 1 - aminopropylagmatine (catalyzed by aminopropyl transferase), and hydrolysis of N 1 -aminopropylagmatine to spermidine by N 1 -aminopropylagmatine ureohydrolase. It is noteworthy that thermophiles synthesize spermidine without producing putrescine as an intermediate. Spermidine can be modified to produce further polyamides such as N 4 -aminopropylspermidine [3(3)4] and then N 4 -bis(aminopropyl) spermidine by an enzyme coded by the TK1691 gene in T. kodakarensis. TK1691 and its orthologues are found in (hyper)thermophilic Archaea and Bacteria, but not in mesophilic Bacteria. TK1691 is a recently characterized aminopropyl transferase involved in the synthesis of branched polyamines, which are essential for the stabilization and structural protection of nucleic acids and which enhance polypeptide synthesis at high temperature.

    Springer Japan, 2015年01月01日, Polyamines: A Universal Molecular Nexus for Growth, Survival, and Specialized Metabolism, 15 - 26, 英語

    [査読有り]

    論文集(書籍)内論文

  • Shinsuke Fujiwara, Ryota Hidese, Takahiro Inoue, Wakao Fukuda

    Gene expression at both the transcriptional and translational levels is critically dependent upon DNA and RNA structure, particularly in hyperthermophiles, which grow at temperatures above 80 °C. Nucleosome-like structures (histone- bound DNA) from hyperthermophilic Archaea are compacted and stabilized in the presence of multivalent polyamines, suggesting that polyamines play a role in nucleosome maintenance in hyperthermophiles. Multivalent polyamines inhibit the melting of double-stranded DNA and structured RNA. Longer-chain polyamines stabilize double-stranded nucleic acids, whereas branched-chain polyamines stabilize stem-and-loop structures, suggesting that branched-chain polyamines are involved in gene translation. Protein synthesis catalyzed by a cell-free extract of the hyperthermophilic archaeon, Thermococcus kodakarensis, requires the presence of longer- and/or branched-chain polyamines. Translational activity increases in the presence of a variety of linear polyamines and is dependent on chain length. Putrescine and spermidine do not increase translational activity. By contrast, longer polyamines such as homocaldopentamine [3334], caldopentamine [3333], and thermopentamine [3343] increase translational activity. The greatest activity occurs in the presence of N 4 -bis(aminopropyl)spermidine [3(3)(3)4] (abbreviation for the number of methylene CH 2 chain units between NH 2, NH, N, or N +). In vitro experiments using cell extracts from the thermophilic bacterium, Thermus thermophiles, reveal that branched-chain polyamines appear to play a role in peptide bond formation during protein biosynthesis. Thus, it appears that branched-chain polyamines are essential for the proper formation of the 30S initiation complex, which acts as the initial aminoacyl-tRNA in thermophiles.

    Springer Japan, 2015年01月01日, Polyamines: A Universal Molecular Nexus for Growth, Survival, and Specialized Metabolism, 143 - 152, 英語

    [査読有り]

    論文集(書籍)内論文

  • Naoki Akasaka, Yuri Ishii, Ryota Hidese, Hisao Sakoda, Shinsuke Fujiwara

    Vinegar with increased amounts of branched-chain amino acids (BCAAs; valine, leucine and isoleucine) is favorable for human health as BCAAs decrease diet-induced obesity and hyperglycemia. To construct Gluconacetobacter europaeus which produces BCAAs, leucine responsive regulator (GeLrp) is focused and two Gelrp mutants were constructed. Wildtype KGMA0119 didn't produce significant amount of valine (0.13 mM) and leucine (0 mM) and strain KGMA7110 which lacks complete Gelrp accumulated valine (0.48 mM) and leucine (0.11 mM) but showed impaired growth, and it was fully restored in the presence of essential amino acids. Strain KGMA7203 was then constructed with a nonsense mutation at codon Trp132 in the Gelrp, which leads a specific deletion at an estimated ligand-sensing region in the C-terminal domain. KGMA7203 produced greater quantities of valine (0.80 mM) and leucine (0.26 mM) and showed the same growth characteristics as KGMA0119. mRNA levels of BCAAs biosynthesis genes (ilvI and ilvC) and probable BCAAs efflux pump (leuE) were determined by quantitative reverse-transcription PCR. Expression rates of ilvI and ilvC in the two Gelrp disruptants were greater than those in KGMA0119. leuE was highly expressed in KGMA7110 only, suggesting that the accumulation in KGMA7110 culture was caused by increased expression of the biosynthesis genes and abnormal enhanced export of amino acids resulting in impaired cell growth. In contrast, KGMA7203 would achieve the high level production through enhanced expression of the biosynthesis genes without enhancing that for the efflux pump. KGMA7203 was considered advantageous for production of vinegar with higher amounts of valine and leucine. (C) 2014, The Society for Biotechnology, Japan. All rights reserved.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2014年12月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 118 (6), 607 - 615, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Ryota Hidese, Takahiro Inoue, Tadayuki Imanaka, Shinsuke Fujiwara

    The sulphur atoms of sulphur-containing cofactors that are essential for numerous cellular functions in living organisms originate from L-cysteine via cysteine desulphurase (CSD) activity. However, many (hyper)thermophilic archaea, which thrive in solfataric fields and are positioned near the root of the evolutionary tree of life, lack CSD orthologues. The existence of CSD orthologues in a subset of (hyper)thermophilic archaea is of interest with respect to the evolution of sulphur-trafficking systems for the cofactors. This study demonstrates that the disruption of the csd gene of Thermococcus kodakarensis, a facultative elemental sulphur (S-0)-reducing hyperthermophilic archaeon, encoding Tk-CSD, conferred a growth defect evident only in the absence of S-0, and that growth can be restored by the addition of S-0, but not sulphide. We show that the csd gene is not required for biosynthesis of thiamine pyrophosphate or molybdopterin, irrespective of the presence or absence of S-0, but is necessary for iron-sulphur cluster biosynthesis in the absence of S-0. Recombinant form of Tk-CSD expressed in Escherichia coli was obtained and it was found to catalyse the desulphuration of L-cysteine. The obtained data suggest that hyperthermophiles might benefit from a capacity for CSD-dependent iron-sulphur cluster biogenesis, which allows them to thrive outside solfataric environments.

    WILEY-BLACKWELL, 2014年07月, MOLECULAR MICROBIOLOGY, 93 (2), 331 - 345, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Kazuma Okada, Ryota Hidese, Wakao Fukuda, Masaru Niitsu, Koichi Takao, Yuhei Horai, Naoki Umezawa, Tsunehiko Higuchi, Tairo Oshima, Yuko Yoshikawa, Tadayuki Imanaka, Shinsuke Fujiwara

    Longer- and/or branched-chain polyamines are unique polycations found in thermophiles. N-4-aminopropylspermine is considered a major polyamine in Thermococcus kodakarensis. To determine whether a quaternary branched penta-amine, N-4-bis(aminopropyl) spermidine, an isomer of N-4-aminopropylspermine, was also present, acid-extracted cytoplasmic polyamines were analyzed by high-pressure liquid chromatography, gas chromatography (HPLC), and gas chromatography-mass spectrometry. N-4-bis(aminopropyl) spermidine was an abundant cytoplasmic polyamine in this species. To identify the enzyme that catalyzes N-4-bis(aminopropyl) spermidine synthesis, the active fraction was concentrated from the cytoplasm and analyzed by linear ion trap-time of flight mass spectrometry with an electrospray ionization instrument after analysis by the MASCOT database. TK0545, TK0548, TK0967, and TK1691 were identified as candidate enzymes, and the corresponding genes were individually cloned and expressed in Escherichia coli. Recombinant forms were purified, and their N-4-bis(aminopropyl) spermidine synthesis activity was measured. Of the four candidates, TK1691 (BpsA) was found to synthesize N4-bis(aminopropyl) spermidine from spermidine via N-4-aminopropylspermidine. Compared to the wild type, the bpsA-disrupted strain DBP1 grew at 85 degrees C with a slightly longer lag phase but was unable to grow at 93 degrees C. HPLC analysis showed that both N-4-aminopropylspermidine and N-4-bis(aminopropyl) spermidine were absent from the DBP1 strain grown at 85 degrees C, demonstrating that the branched-chain polyamine synthesized by BpsA is important for cell growth at 93 degrees C. Sequence comparison to orthologs from various microorganisms indicated that BpsA differed from other known aminopropyltransferases that produce spermidine and spermine. BpsA orthologs were found only in thermophiles, both in archaea and bacteria, but were absent from mesophiles. These findings indicate that BpsA is a novel aminopropyltransferase essential for the synthesis of branched-chain polyamines, enabling thermophiles to grow in high-temperature environments.

    AMER SOC MICROBIOLOGY, 2014年05月, JOURNAL OF BACTERIOLOGY, 196 (10), 1866 - 1876, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Ryota Hidese, Ryo Nishikawa, Le Gao, Masahiro Katano, Tomohiro Imai, Satoru Kato, Tamotsu Kanai, Haruyuki Atomi, Tadayuki Imanaka, Shinsuke Fujiwara

    Two genes, TK1280 and TK2287, encode orthologous transcription factor B proteins (TFB1 and TFB2, respectively) in the hyperthermophilic archaeon Thermococcus kodakarensis. The functional difference between their TFBs remains unknown. While TFB1 and TFB2 displayed equivalent thermostability, mRNA levels of tfb1 at 93 A degrees C were eightfold higher than those at 60 or 85 A degrees C, and were 4- to 10-fold greater than those of tfb2 at all temperatures. This suggests that TFB1 is the abundant TFB in T. kodakarensis and is heat-inducible. By contrast, the mRNA level of tfb2 increased at 93 A degrees C, but the levels were less than twofold of those at 60 or 85 A degrees C. No significant differences in growth were observed among the DTF1 (a dagger tfb1, a dagger pyrF), DTF2 (a dagger tfb2 a dagger pyrF), and parental host strain KU216 (a dagger pyrF) at 60 A degrees C. However, DTF2 showed a decrease in cell yield at 85 A degrees C, and both DTF1 and DTF2 showed growth defects at 93 A degrees C. Comparative transcriptome analysis between KU216 and DTF1 or DTF2 indicated that TFB1 apparently controls the expression of genes essential for motility/adhesion, whereas TFB2 regulates genes involved in mevalonate/lipid biosynthesis. In DTF1, the ratio of cells with flagella decreased at 85 and 93 A degrees C, and reporter studies indicated that flaB1 transcription is dependent on TFB1 at 85 A degrees C but not at 60 A degrees C.

    SPRINGER JAPAN KK, 2014年05月, EXTREMOPHILES, 18 (3), 573 - 588, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Ryota Hidese, Hisaaki Mihara, Tatsuo Kurihara, Nobuyoshi Esaki

    Iron-sulfur (Fe-S) clusters are ubiquitous cofactors that are crucial for many physiological processes in all organisms. In Escherichia coli, assembly of Fe-S clusters depends on the activity of the iron-sulfur cluster (ISC) assembly and sulfur mobilization (SUF) apparatus. However, the underlying molecular mechanisms and the mechanisms that control Fe-S cluster biogenesis and iron homeostasis are still poorly defined. In this study, we performed a global screen to identify the factors affecting Fe-S cluster biogenesis and iron homeostasis using the Keio collection, which is a library of 3,815 single-gene E. coli knockout mutants. The approach was based on radiolabeling of the cells with [2-C-14] dihydrouracil, which entirely depends on the activity of an Fe-S enzyme, dihydropyrimidine dehydrogenase. We identified 49 genes affecting Fe-S cluster biogenesis and/or iron homeostasis, including 23 genes important only under microaerobic/anaerobic conditions. This study defines key proteins associated with Fe-S cluster biogenesis and iron homeostasis, which will aid further understanding of the cellular mechanisms that coordinate the processes. In addition, we applied the [2-C-14] dihydrouracil-labeling method to analyze the role of amino acid residues of an Fe-S cluster assembly scaffold (IscU) as a model of the Fe-S cluster assembly apparatus. The analysis showed that Cys37, Cys63, His105, and Cys106 are essential for the function of IscU in vivo, demonstrating the potential of the method to investigate in vivo function of proteins involved in Fe-S cluster assembly.

    AMER SOC MICROBIOLOGY, 2014年03月, JOURNAL OF BACTERIOLOGY, 196 (6), 1238 - 1249, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Naoki Akasaka, Hisao Sakoda, Ryota Hidese, Yuri Ishii, Shinsuke Fujiwara

    Gluconacetobacter europaeus, one of the microorganisms most commonly used for vinegar production, produces the unfavorable flavor compound acetoin. Since acetoin reduction is important for rice vinegar production, a genetic approach was attempted to reduce acetoin produced by G. europaeus KGMA0119 using specific gene knockout without introducing exogenous antibiotic resistance genes. A uracil-auxotrophic mutant with deletion of the orotate phosphoribosyltransferase gene (pyrE) was first isolated by positive selection using 5-fluoroorotic acid. The pyrE disruptant designated KGMA0704 (Delta pyrE) showed 5-fluoroorotic acid resistance. KGMA0704 and the pyrE gene were used for further gene disruption experiments as a host cell and a selectable marker, respectively. Targeted disruption of aldC or als, which encodes alpha-acetolactate decarboxylase or alpha-acetolactate synthase, was attempted in KGMA0704. The disruption of these genes was expected to result in a decrease in acetoin levels. A disruption vector harboring the pyrE marker within the targeted gene was constructed for double-crossover recombination. The cells of KGMA0704 were transformed with the exogenous DNA using electroporation, and genotypic analyses of the transformants revealed the unique occurrence of targeted aldC or als gene disruption. The aldC disruptant KGMA4004 and the als disruptant KGMA5315 were cultivated, and the amount of acetoin was monitored. The acetoin level in KGMA4004 culture was significantly reduced to 0.009% (wt/vol) compared with KGMA0119 (0.042% [wt/vol]), whereas that of KGMA5315 was not affected (0.037% [wt/vol]). This indicates that aldC disruption is critical for acetoin reduction. G. europaeus KGMA4004 has clear application potential in the production of rice vinegar with less unfavorable flavor.

    AMER SOC MICROBIOLOGY, 2013年12月, APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 79 (23), 7334 - 7342, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Eriko Nagaoka, Ryota Hidese, Tadayuki Imanaka, Shinsuke Fujiwara

    Thermococcus kodakarensis, which grows optimally at 85 degrees C, expresses cold stress-inducible DEAD box RNA helicase (Tk-deaD) when shifted to 60 degrees C. A DDA1 deletion (Delta Tk-deaD) mutant exhibited decreased cell growth, and cells underwent lysis at 60 degrees C in nutrient broth in the absence of elemental sulfur. In contrast, cells in medium containing elemental sulfur at 60 degrees C did not undergo lysis, suggesting that Tk-deaD is necessary for cell growth in sulfur-free medium. To identify the element responsible for the cold response, a pTKR expression probe plasmid was constructed using thermostable catalase from Pyrobaculum calidifontis as a reporter. The plasmid pTKRD, which contained the transcription factor B recognition element, TATA region, and Shine-Dalgarno (SD) region, including the initiation codon of the Tk-deaD gene, exhibited cold inducibility. We also constructed a series of deletion and chimeric constructs with the glutamate dehydrogenase (gdh) promoter, whose expression is constitutive independent of culture temperatures and catalase expression. Reporter assay experiments indicated that the regulatory element is located in the region between the SD region and the initiation codon (ATG). Nucleotide sequences in the upstream regions of Tk-deaD and gdh were compared and revealed a five-adenosine (AAAAA) sequence between SD and ATG of Tk-deaD that was not present in gdh. Replacement of the repeated adenosine sequence with other sequences revealed that the AAAAA sequence is important for cold induction. This sequence-specific mechanism is unique and is one that has not been identified in other known cold-inducible genes.

    AMER SOC MICROBIOLOGY, 2013年08月, JOURNAL OF BACTERIOLOGY, 195 (15), 3442 - 3450, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Ryota Hidese, Hisaaki Mihara, Tatsuo Kurihara, Nobuyoshi Esaki

    The pyrimidine reductive catabolic pathway is important for the utilization of uracil and thymine as sources of nitrogen and carbon. The pathway is controlled by three enzymes: dihydropyrimidine dehydrogenase (DPD), dihydropyrimidinase and beta-alanine synthase. The putative DPD genes, pydX and pydA, are tandemly arranged in the Pseudomonas putida genome. Intriguingly, a putative transcriptional regulator, PydR, homologous to Escherichia coli RutR, a repressor of the Rut-dependent pyrimidine degradation pathway, is located downstream of pydX and pydA. In this study, we show that a pydA strain of P. putida fails to grow on a minimal media containing uracil or thymine as a sole nitrogen source, demonstrating the physiological importance of DPD in the reductive pathway. The expression of pydA and DPD activity in the absence of uracil were significantly higher in a pydR strain than in the wild-type strain, indicating that PydR acts as a repressor of the pyrimidine reductive pathway in P. putida. Phylogenetic analysis of RutR and PydR suggests that these homologous repressors may have evolved from a common ancestral protein that regulates pyrimidine degradation.

    OXFORD UNIV PRESS, 2012年10月, JOURNAL OF BIOCHEMISTRY, 152 (4), 341 - 346, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Ryota Hidese, Hisaaki Mihara, Nobuyoshi Esaki

    Cysteine desulfurases are pyridoxal 5'-phosphate-dependent homodimeric enzymes that catalyze the conversion of L-cysteine to L-alanine and sulfane sulfur via the formation of a protein-bound cysteine persulfide intermediate on a conserved cysteine residue. The enzymes are capable of donating the persulfide sulfur atoms to a variety of biosynthetic pathways for sulfur-containing biofactors, such as iron-sulfur clusters, thiamin, transfer RNA thionucleosides, biotin, and lipoic acid. The enormous advances in biochemical and structural studies of these biosynthetic pathways over the past decades provide an opportunity for detailed understanding of the nature of the excellent sulfur transfer mechanism of cysteine desulfurases.

    SPRINGER, 2011年07月, APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 91 (1), 47 - 61, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Ryota Hidese, Hisaaki Mihara, Tatsuo Kurihara, Nobuyoshi Esaki

    The reductive pyrimidine catabolic pathway is absent in Escherichia coli. However, the bacterium contains an enzyme homologous to mammalian dihydropyrimidine dehydrogenase. Here, we show that E. coli dihydropyrimidine dehydrogenase is the first member of a novel NADH-dependent subclass of iron-sulfur flavoenzymes catalyzing the conversion of uracil to 5,6-dihydrouracil in vivo.

    AMER SOC MICROBIOLOGY, 2011年02月, JOURNAL OF BACTERIOLOGY, 193 (4), 989 - 993, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Functional analysis of a novel NADH-dependent dihydropyrimidine dehydrogenase from Escherichia coli

    HIDESE Ryota, MIHARA Hisaaki, ESAKI Nobuyoshi

    2009年09月, Vitamins, 83 (9), 528 - 532, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Hisaaki Mihara, Ryota Hidese, Masahiro Yamane, Tatsuo Kurihara, Nobuyoshi Esaki

    inactivation of iscS encoding cysteine desulfurase results in a slow growth phenotype associated with the deficiency of iron-sulfur clusters, thiamine, NAD, and tRNA thionucleosides in Escherichia coli. However, the other roles of iscS in vivo are unknown. By using differential screening strategies, we identified 2 pyrimidine salvage enzymes, namely, uridine phosphorylase and cytidine deaminase, which were down-regulated in the iscS mutant. Both enzymes are positively regulated by the cAMP receptor protein (CRP). We also identified a novel protein complex, namely, YeiT-YeiA, whose expression level was decreased in the iscS mutant. The recombinant YeiT-YeiA complex exhibited NADH-dependent dihydropyrimidine dehydrogenase activity, indicating its role in pyrimidine metabolism. The presence of a CRP-binding consensus sequence on the 5'-upstream of the yeiT-YeiA gene suggests its regulation by CRP. These results provide a clue to the possible role of iscS in pyrimidine metabolism by gene regulation. (C) 2008 Elsevier Inc. All rights reserved.

    ACADEMIC PRESS INC ELSEVIER SCIENCE, 2008年08月, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 372 (3), 407 - 411, 英語

    [査読有り]

    研究論文(学術雑誌)

MISC

  • 耐熱性ヘリカーゼを利用した高精度核酸検出技術の開発

    藤原 伸介, 藤原 綾子, 秀瀨 涼太

    2017年07月, 生物工学会誌, 95 (7), 383 - 387, 日本語

    記事・総説・解説・論説等(学術雑誌)

  • 超好熱性アーキアThermococcus kodakarensis由来分岐鎖ポリアミン合成酵素の構造と機能

    秀瀨 涼太, 藤原 伸介

    2016年04月, 酵素工学ニュース, 75, 日本語

    記事・総説・解説・論説等(学術雑誌)

  • 進化は長い目で見て

    秀瀨 涼太

    2016年03月, 生物工学会誌, 94 (3), 131, 日本語

    記事・総説・解説・論説等(学術雑誌)

  • 好熱菌の低温ストレス耐性

    秀瀨 涼太, 藤原 伸介

    2015年08月, 生物工学会誌, 93 (8), 464 - 467, 日本語

    記事・総説・解説・論説等(学術雑誌)

  • 耐熱性タンパク質を利用したビスフェノールAの吸着

    秀瀨 涼太, 今岡 進, 藤原 伸介

    2014年04月, ケミカルエンジニヤリング, 59 (4), 17 - 21, 日本語

    記事・総説・解説・論説等(学術雑誌)

  • 高温において翻訳活性化作用を持つポリアミン

    秀瀨 涼太, 藤原 伸介

    2012年05月, バイオサイエンスとインダストリー, 70 (3), 211 - 212, 日本語

    記事・総説・解説・論説等(学術雑誌)

  • シャペロニンと生物進化

    秀瀨 涼太

    2012年01月, 生物工学会誌, 90 (1), 39, 日本語

    記事・総説・解説・論説等(学術雑誌)

書籍等出版物

  • 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 「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

    共著, シーエムシー出版, 2018年06月, 日本語

    学術書

  • Identification of branched-chain polyamines in hyperthermophiles, In Methods in Molecular Biology

    HIDESE Ryota, FUKUDA Wakao, NIITSU Masaru, FUJIWARA Shinsuke

    共著, Springer, 2018年, 英語

    学術書

  • Protein synthesis and polyamines in thermophiles: Effect of polyamines on nucleic acid maintenance and gene expression, In Polyamines, a universal molecular nexus for growth, survival and specialized metabolism

    FUJIWARA Shinsuke, HIDESE Ryota, INOUE Takahiro, FUKUDA Wakao

    共著, Springer, 2015年11月, 英語

    学術書

  • Long-chain and branched polyamines in thermophilic microbes. In Polyamines, a universal molecular nexus for growth, survival and specialized metabolism

    FUKUDA Wakao, HIDESE Ryota, FUJIWARA Shinsuke

    共著, Springer, 2015年, 英語

    学術書

講演・口頭発表等

  • Physiological roles of Thermococcus kodakarensis ubiquitin-like proteins in the biosynthesis of sulfur-containing biomolecules.

    Ryota Hidese, Satsuki Sakakibara, Takayuki Ohira, Tsutomu Suzuki, Naoki Shigi, Shinsuke Fujiwara

    Thermophiles2019, 2019年09月, 英語, 国際会議

    ポスター発表

  • 分岐鎖ポリアミンが超好熱菌のRNA ポリメラーゼ複合体に及ぼす影響

    福田 萌子, 家森 優佳, 濱川 匡史, 秀瀨 涼太, 福田 青郎, 藤原 伸介

    第71回日本生物工学会大会, 2019年09月, 日本語, 岡山大学, 国内会議

    口頭発表(一般)

  • カチオンの構造に依存した負電荷脂質膜の相分離

    永田 佳嗣, 引地 啓太, 秀瀨 涼太, 藤原 伸介, 下川 直史, 高木 昌宏

    第71回日本生物工学会大会, 2019年09月, 日本語, 岡山大学, 国内会議

    口頭発表(一般)

  • 微生物代謝経路の解析と生物生産への応用

    秀瀨 涼太

    第5回 生物資源セミナー, 2019年01月, 日本語, 立命館大学びわこくさつキャンパス, 国内会議

    口頭発表(一般)

  • 硫黄転移関連タンパク質から紐解く生命の進化

    秀瀨 涼太

    極限環境生物学会2018年度(第19回)年会, 2018年12月, 日本語, 松江コンベンションセンター, 国内会議

    [招待有り]

    口頭発表(招待・特別)

  • 超好熱性アーキアにおけるユビキチン様タンパク質の機能解析

    榊原 早津季, 秀瀨 涼太, 藤原 伸介

    第70回日本生物工学会大会, 2018年09月, 日本語, 関西大学 千里山キャンパス, 国内会議

    ポスター発表

  • ハイスループット微生物構築・評価技術の開発

    石井 純, 西 晶子, 北野 美保, 中村 朋美, 加藤 寛子, 庄司 信一郎, 秀瀨 涼太, 梅野 太輔

    第20回 新産業技術促進検討会, 2018年07月, 日本語, モノづくり日本会議/日刊工業新聞社, 国内会議

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