Elucidation of the maturation mechanism of subtilisins from hyperthermophiles and development of their potential use

2011 Fiscal Year Final Research Report

• Project/Area Number: 21380065
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Applied biochemistry
• Research Institution: Osaka University
• Principal Investigator: KANAYA Shigenori 大阪大学, 大学院・工学研究科, 教授 (30273585)
• Co-Investigator(Renkei-kenkyūsha): KOGA Yuichi 大阪大学, 大学院・工学研究科, 准教授 (30379119)
• Project Period (FY): 2009 – 2011
• Keywords: タンパク質工学 / 微生物酵素 / 構造生物学

Research Abstract

The maturation mechanisms of subtilisins, Tk-subtilisin and Tk-SP, from the hyperthermophilic archaeon Thermococcus kodakarensis were analyzed. The results indicate that both proteins are activated(matured) upon autoprocessing and degradation of N-terminal propeprides, folding of Tk-subtilisin is induced upon binding of the Ca^<2+> ions to the Ca^<2+>-binding loop, Tk-SP is matured in the absence of the Ca^<2+> ions, Tk-SP requires C-terminalβ-jelly roll domain for hyperstability. It was also shown that Tk-subtilisin is useful for degradation of abnormal prion proteins.

Research Products

• [Journal Article] An alternative mature form of subtilisin homologue, Tk-SP, from Thermococcus kodakaraensis identified in the presence of Ca^<2+> (2011)
  • Author(s): Sinsereekul, N., Foophow, T., Yamanouchi, M., Koga, Y., Takano, K. and Kanaya, S.
  • Journal Title: FEBS J Volume: 278 Pages: 1901-1911
  • URL: http://www.ncbi.nlm.nih.gov/pubmed/2144
  • Peer Reviewed
• [Journal Article] Inhibition of chymotrypsin-and subtilisin-like serine proteases with Tk-serpin from hyperthermophilic archaeon Thermococcu skodakaraensis (2011)
  • Author(s): Tanaka, S., Koga, Y., Takano, K., and Kanaya, S.
  • Journal Title: Biochim. Biophys. Acta Volume: 1814 Pages: 299-307
  • URL: http://www.ncbi.nlm.nih.gov/pubmed/2111
  • Peer Reviewed
• [Journal Article] Crystal structure of a subtilisin homologue, Tk-SP, from Thermococcus kodakaraensis : requirement of a C-terminalβ-jelly roll domain for hyperstability (2010)
  • Author(s): Foophow, T., Tanaka, S., Angkawidjaja, C., Koga, Y., Takano, K., and Kanaya, S.
  • Journal Title: J. Mol. Biol Volume: 400 Pages: 865-877
  • URL: http://www.ncbi.nlm.nih.gov/pubmed/20595040
  • Peer Reviewed
• [Journal Article] Identification of the interactions critical for propeptide-catalyzed folding of Tk-subtilisin (2009)
  • Author(s): Tanaka, S., Matsumura, H., Koga, Y., Takano, K., and Kanaya, S.
  • Journal Title: J. Mol. Biol Volume: 394 Pages: 306-319
  • URL: http://www.ncbi.nlm.nih.gov/pubmed/19766655
  • Peer Reviewed
• [Journal Article] Requirement of a unique Ca^<2+>-binding loop for folding of Tk-subtilisin from a hyperthermophilic archaeon (2009)
  • Author(s): Takeuchi, Y., Tanaka, S., Matsumura, H., Koga, Y., Takano, K., and Kanaya, S.
  • Journal Title: Biochemistry Volume: 48 Pages: 10637-10643
  • URL: http://www.ncbi.nlm.nih.gov/pubmed/19813760
  • Peer Reviewed
• [Presentation] Unique maturation mechanism of Tk-subtilisin from a hyperthermophilic archaeon (2012)
  • Author(s): 金谷茂則
  • Organizer: 2011年度日本農芸化学会(シンポジウム)
  • Place of Presentation: 京都女子大学(京都)
  • Year and Date: 20120322-25
• [Presentation] Unique folding and stabilization mechanisms of subtilisin-like serine proteases from a hyperthermophilic archaeon (2011)
  • Author(s): Shigenori Kanaya
  • Organizer: International Conference on Enzyme Science and Technology
  • Place of Presentation: Kusadasi(Turkey)
  • Year and Date: 20111031-1104
• [Presentation] 超好熱菌由来プロテアーゼによる異常プリオン蛋白質分解 (2010)
  • Author(s): 古賀雄一、田中俊一、作道章一、高野和文、金谷茂則
  • Organizer: 日本生物工学会
  • Place of Presentation: フェニックス・シーガイア・リゾート(宮崎)
  • Year and Date: 20101027-29
• [Presentation] Degradation of abnormal prion protein by a new protease from a hyperthermophile (2010)
  • Author(s): Yuichi Koga, Shun-ichi Tanaka, Akikazu Sakudo, Kazuyoshi Ikuta, Kazufumi Takano, and Shigenori Kanaya.
  • Organizer: PRION 2010
  • Place of Presentation: Salzburg(Austria)
  • Year and Date: 20100908-11
• [Presentation] 高アルカリ性、高温でのTk-subtilisinの成熟化促進機構の解析 (2009)
  • Author(s): 上田泰徳、田中俊一、古賀雄一、高野和文、金谷茂則
  • Organizer: 日本生化学会
  • Place of Presentation: 神戸国際会議場(神戸市)
  • Year and Date: 20091020-24
• [Patent(Industrial Property Rights)] プロテアーゼの製造方法、並びにプロテアーゼ溶液およびプロテアーゼのプロ体溶液 (2009)
  • Inventor(s): 金谷茂則、田中俊一、竹内勇希、高野和文、古賀雄一
  • Industrial Property Rights Holder: 大阪大学
  • Industrial Property Number: 特許, PCT/ JP2009/ 64629
  • Filing Date: 2009-08-21
  • Overseas
• [Patent(Industrial Property Rights)] 新規なプロテアーゼおよびその利用 (2009)
  • Inventor(s): 金谷茂則、チタフーパオ、高野和文、古賀雄一
  • Industrial Property Rights Holder: 大阪大学
  • Industrial Property Number: 特許, PCT/ JP2009/ 063547
  • Filing Date: 2009-07-30