Studies on the enzymes related to central metabolism of hyperthermophilic archaea

• Project/Area Number: 24580136
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Applied biochemistry
• Research Institution: The University of Tokyo
• Principal Investigator: WAKAGI Takayoshi 東京大学, 農学生命科学研究科, 研究員 (70175058)
• Project Period (FY): 2012-04-01 – 2015-03-31
• Project Status: Completed (Fiscal Year 2014)
• Budget Amount: ¥5,070,000 (Direct Cost: ¥3,900,000、Indirect Cost: ¥1,170,000); Fiscal Year 2014: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000); Fiscal Year 2013: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000); Fiscal Year 2012: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
• Keywords: 酵素 / 酸化還元酵素 / 古細菌 / 超好熱菌 / フェレドキシン / アロステリズム / 中央代謝 / 結晶構造解析 / モリブデン酵素 / アロステリック酵素

Outline of Final Research Achievements

Structure-function relationships of several enzymes related to central metabolism of acidophilic and hyperthermophilic archaeon were investigated. Glyceraldehyde-3-phosphate dehydrogenases including traditional phosphorylating type GAPDH and non-phosphrylating enzyme GAPN were characterized. Single amino acid substitution of GAPN resulted in conversion of the allosterism type from K into V.
Three isozymes of glyceraldehyde oxidoreductase related to glycolysis were isolated. They showed novel subunit compositions and the crystal structure suggested characteristic molecular construction pathway.
2-Oxoacid:ferredoxin oxidoreductase from the organism has only one iron-sulfur clusted ligated by 4 Cys. Substitution of Cys and X-ray crystallography suggested a novel reaction mechanism. Ferredoxin NADP(+) reductase was identified by analyzing ST2133.

Research Products

• [Journal Article] Sulfolobus tokodaii ST2133 is characterized as a thioredoxin reductase-like ferredoxin:NADP+ oxidoreductase (2014)
  • Author(s): Z. Yan, Y.-W. Nam, S. Fushinobu, and T. Wakagi
  • Journal Title: Extremophiles Volume: 18 Issue: 1 Pages: 99-110
  • DOI: 10.1007/s00792-013-0601-1
  • Peer Reviewed
• [Journal Article] Four Cys residues in heterodimeric 2-oxoacid:ferredoxin oxidoreductase are required for CoA-dependent oxidative decarboxylation but not for a non-oxidative decarboxylation (2014)
  • Author(s): Z. Yan, S. Fushinobu, and T. Wakagi
  • Journal Title: Biochim. Biophys. Acta Volume: 1844 Issue: 4 Pages: 736-743
  • DOI: 10.1016/j.bbapap.2014.01.015
  • Peer Reviewed
• [Journal Article] Engineering the allosteric properties of archaeal non-phosphorylating glyceraldehyde-3-phosphate dehydrogenases. (2014)
  • Author(s): Fumiaki Ito, Masayuki Miyake, Shinya Fushinobu, Shugo Nakamura, Kentaro Shimizu, Takayoshi Wakagi
  • Journal Title: BBA - Proteins and Proteomics Volume: Volume 1844 Issue: 4 Pages: 759-766
  • DOI: 10.1016/j.bbapap.2014.01.017
  • Peer Reviewed / Open Access
• [Journal Article] Aldolase-Distinctive Characters in Archaeal Enzymes (2013)
  • Author(s): Takayoshi Wakagi
  • Journal Title: Trends in Glycoscience and Glycotechnology Volume: 25 Issue: 142 Pages: 71-81
  • DOI: 10.4052/tigg.25.71
  • NAID: 10031190621
  • ISSN: 0915-7352, 1883-2113
  • Peer Reviewed
• [Journal Article] Archaeal Aldehyde Dehydrogenase ST0064 from <i>Sulfolobus tokodaii</i>, a Paralog of Non-Phosphorylating Glyceraldehyde-3-phosphate Dehydrogenase, Is a Succinate Semialdehyde Dehydrogenase (2013)
  • Author(s): F. Ito, H. Chishiki, S. Fushinobu, and T. Wakagi
  • Journal Title: Bioscience, Biotechnology, and Biochemistry Volume: 77 Issue: 6 Pages: 1344-1348
  • DOI: 10.1271/bbb.130119
  • NAID: 10031184711
  • ISSN: 0916-8451, 1347-6947
  • Peer Reviewed
• [Journal Article] アルドラーゼ ー 古細菌酵素の特殊性 (2013)
  • Author(s): 若木高善
  • Journal Title: Trends in Glycoscience and Glycotechnology Volume: in press
  • NAID: 10031190621
  • Peer Reviewed
• [Journal Article] Comparative analysis of two glyceraldehyde-3-phosphate dehydrogenases from a thermoacidophilic archaeon, Sulfolobus tokodaii (2012)
  • Author(s): Fumiaki Ito, Hidehiro Chishiki, Shinya Fushinobu, Takayoshi Wakagi
  • Journal Title: FEBS Letters Volume: 586 Issue: 19 Pages: 3097-3103
  • DOI: 10.1016/j.febslet.2012.07.059
  • Peer Reviewed
• [Journal Article] 「真の」二機能性酵素の発見とその「変身」のメカニズム (2012)
  • Author(s): 伏信進矢、西増弘志、若木高善
  • Journal Title: 化学と生物 Volume: 50 (2) Pages: 868-875
• [Journal Article] 1つの酵素が2つの反応を触媒するしくみ (2012)
  • Author(s): 西増弘志、伏信進矢、若木高善
  • Journal Title: 日本結晶学会誌 Volume: 54 (2) Pages: 113-118
  • NAID: 10030631990
• [Presentation] 超好熱性古細菌フェレドキシンNADP+レダクターゼの構造と機能 (2015)
  • Author(s): Zhen Yan、馬込明音、荒川孝俊、伏信進矢、○若木高善
  • Organizer: ２０１５年度日本農芸化学会年会
  • Place of Presentation: 岡山大学津島キャンパス
  • Year and Date: 2015-03-28
• [Presentation] 超好熱菌の原始的中央代謝に見られるいくつかの興味深い酵素たち (2014)
  • Author(s): 若木高善
  • Organizer: 第３７回日本分子生物学会年会
  • Place of Presentation: 横浜みなとみらい
  • Year and Date: 2014-11-25
  • Invited
• [Presentation] 超好熱性古細菌由来無機ピロリン酸依存性ホスホフルクトキナーゼの 構造と機能 (2014)
  • Author(s): 宋賢珍、日高将文、荒川隆俊、伏信進矢、○若木高善
  • Organizer: 日本極限環境生物学会年会
  • Place of Presentation: 沖縄県名護市
  • Year and Date: 2014-11-03
• [Presentation] 超好熱菌の原始的中央代謝に見られるいくつかの興味深い酵素たち (2014)
  • Author(s): 若木高善
  • Organizer: 千葉工大フォーラム
  • Place of Presentation: 千葉工大
  • Year and Date: 2014-10-28
  • Invited
• [Presentation] 好酸好熱性古細菌Sulfolobus tokodaii由来のnon-phosphorylating glyceraldehyde-3-phosphate dehydrogenaseにおけるアロステリック制御の解析 (2013)
  • Author(s): 伊藤史晃・知識秀裕・中村周吾・清水謙多郎・伏信進矢・若木高善
  • Organizer: 日本農芸化学会大会
  • Place of Presentation: 東北大学
• [Presentation] Ferredoxin:NADP+ oxidoreductase from Sulfolobus Tokodaii (2013)
  • Author(s): Yan Zhen・Nam Yoon-Woo・伏信進矢・若木高善
  • Organizer: 日本農芸化学会大会
  • Place of Presentation: 東北大学
• [Presentation] 好酸好熱性古細菌Sulfolobus tokodaii由来の非リン酸化型グリセルアルデヒド-3-リン酸脱水素酵素におけるアロステリック制御の解析 (2012)
  • Author(s): 伊藤史晃・知識秀祐・中村周吾・清水謙多郎・伏信進矢・若木高善
  • Organizer: 日本極限環境生物学会年会
  • Place of Presentation: 明治大学文理学部
• [Presentation] Study on the catalytic reaction mechanism of 2-oxoacid:ferredoxin oxidoreductase from Sulfolobus tokodaii (2012)
  • Author(s): Yan Zhen・伏信進矢・若木高善
  • Organizer: 日本極限環境生物学会年会
  • Place of Presentation: 明治大学文理学部
• [Presentation] 好酸好熱性古細菌i由来の非リン酸化型グリセルアルデヒド-3-リン酸脱水素酵素におけるアロステリック制御の解析 (2012)
  • Author(s): 伊藤史晃・知識秀祐・中村周吾・清水謙多郎・伏信進矢・若木高善
  • Organizer: 日本農芸化学会関東支部大会
  • Place of Presentation: 新潟薬科大学
• [Presentation] Role of iron-sulfur cluster on the dual activities of 2-oxoacid:ferredoxin oxidoreductase from Sulfolobus tokodaii (2012)
  • Author(s): Zhen Yan, Shinya Fushinobu, Takayoshi Wakagi
  • Organizer: The 12th Japan-China-Korea Joint Symposium on Enzyme Engineering
  • Place of Presentation: Kanazawa Excel Hotel Tokyu
• [Presentation] Conversion of an iron sulfur cluster in heterodimeric 2-oxoacid:ferredoxin oxidoreductase from Sulfolobus tokodaii
  • Author(s): Zhen Yan, Shinya Fushinobu, and ○Takayoshi Wakagi
  • Organizer: 日本極限環境生物学会年会
  • Place of Presentation: 明治大学生田校舎
• [Presentation] Intact iron-sulfur cluster is required for oxidative decarboxylation of pyruvate:ferredoxin oxidoreductase from Sulfolobus tokodaii
  • Author(s): Zhen Yan, Shinya Fushinobu, and ○Takayoshi Wakagi
  • Organizer: 第１３回日本蛋白質科学会年会
  • Place of Presentation: とりぎん文化会館
• [Presentation] 好酸好熱性古細菌Sulfolobus tokodaii 由来の非リン酸化型グリセルアルデヒド-3-リン酸脱水素酵素におけるアロステリック制御にともなう構造変化
  • Author(s): 伊藤史晃、三宅将之、中村周吾、清水 謙多郎、伏信進矢、○若木高善
  • Organizer: 第１３回日本農芸化学会年会
  • Place of Presentation: 明治大学生田校舎
• [Presentation] 超好熱性微生物由来の無機ピロリン酸依存性ホスホフルクトキナーゼの結晶構造解析とリン酸供与体の認識機構
  • Author(s): ○宋賢珍、日高将文、荒川隆俊、伏信進矢、若木高善
  • Organizer: ２０１４年度日本農芸化学会大会
  • Place of Presentation: 明治大学生田校舎