Biosynthetic study on bioactive compounds from marine sponges

• Project/Area Number: 17F17114
• Research Category: Grant-in-Aid for JSPS Fellows
• Allocation Type: Single-year Grants
• Section: 外国
• Research Field: Natural medicines
• Research Institution: Hokkaido University
• Principal Investigator: 脇本 敏幸 北海道大学, 薬学研究院, 教授 (70363900)
• Co-Investigator(Kenkyū-buntansha): URIA AGUSTINUS 北海道大学, 薬学研究科(研究院), 外国人特別研究員
• Project Period (FY): 2017-07-26 – 2019-03-31
• Project Status: Completed (Fiscal Year 2018)
• Budget Amount: ¥2,400,000 (Direct Cost: ¥2,400,000); Fiscal Year 2018: ¥1,200,000 (Direct Cost: ¥1,200,000); Fiscal Year 2017: ¥1,200,000 (Direct Cost: ¥1,200,000)
• Keywords: 海綿 / メタゲノム / 生合成

Outline of Annual Research Achievements

海綿動物は多種多様な共生微生物を宿す付着性の多細胞動物である。世界中の海に生息し、１万種が現存するといわれる海綿動物からは、これまで２万以上の生理活性物質が単離、報告されてきた。ランダムスクリーニングに由来するため細胞毒性物質の数が多く、pM程度の濃度で強力な細胞毒性を示す化合物も少なくない。しかしながら、それらの多くが稀少かつ複雑な構造を有するために、抗がん剤として研究開発がなされた化合物は極めて少ない。一方で、それら海綿由来細胞毒性物質のほとんどは共生微生物によって生産されることが示唆されている。本研究では医薬品資源として有望な海綿由来稀少生理活性物質の生合成遺伝子および生産菌を同定し、可培養化や異種生産を目指した基礎的知見を得る。昨年度より海綿動物から微量成分として見出されたspongistatinの生合成遺伝子の特定を進めてきた。Spongistatinはtrans AT型PKSによる生合成経路が予想される化合物であり、その生合成遺伝子には特徴的な配列を有するketosynthase (KS)が含まれる。したがってKSの縮重プライマーを用いたtrans AT型PKSの探索を重点的に行った。その結果、海綿メタゲノムDNAよりtrans AT型PKSに属するKS配列を得た。さらに海綿メタゲノムライブラリーを構築し、得られたtrans AT型KS配列に基づくスクリーニングを行った。その結果、複数のヒットクローンを得ることに成功した。現在詳細なシークエンス解析を進めている。また、海綿共生微生物の可培養化を目指し、沖縄産海綿Theonella swinhoeiから様々な組成の培地を用いた培養を試みた。その結果、既知微生物と相同性が低い微生物を含む数百種類の微生物を単離した。その中にはPKSやNRPS遺伝子を有する微生物群も数多く含まれており今後さらに物質生産能の検証を進める。

Research Progress Status

平成30年度が最終年度であるため、記入しない。

Strategy for Future Research Activity

平成30年度が最終年度であるため、記入しない。

Research Products

• [Int'l Joint Research] スイス連邦工科大学チューリッヒ校(スイス)
• [Journal Article] SurE is a trans-acting thioesterase cyclizing two distinct non-ribosomal peptides (2019)
  • Author(s): Matsuda, K., Kobayashi, M., Kuranaga, T., Takagda, K., Ikeda, H., Matsunaga, S., Wakimoto, T.,
  • Journal Title: Org. Biomol. Chem. Volume: 17 Issue: 5 Pages: 1058-1061
  • DOI: 10.1039/c8ob02867b
  • Peer Reviewed / Open Access
• [Journal Article] The revised structure of surugamide A (2019)
  • Author(s): Matsuda Kenichi、Kuranaga Takefumi、Ayae Sano, Akihiro Ninomiya、Takada Kentaros、Wakimoto Toshiyuki
  • Journal Title: Chemical and Pharmaceutical Bulletin Volume: -
  • Peer Reviewed
• [Journal Article] Diastereoselective Total Synthesis and Structural Confirmation of Surugamide F (2018)
  • Author(s): Kuranaga Takefumi、Fukuba Atsuki、Ninomiya Akihiro、Takada Kentaro、Matsunaga Shigeki、Wakimoto Toshiyuki
  • Journal Title: Chemical and Pharmaceutical Bulletin Volume: 66 Issue: 6 Pages: 637-641
  • DOI: 10.1248/cpb.c18-00072
  • NAID: 130007382323
  • ISSN: 0009-2363, 1347-5223
  • Year and Date: 2018-06-01
  • Peer Reviewed
• [Journal Article] Biosynthetic Insights of Calyculin- and Misakinolide-Type Compounds in “ Candidatus Entotheonella sp.” (2018)
  • Author(s): Uria Agustinus R.、Piel Joern、Wakimoto Toshiyuki
  • Journal Title: Methods in Enzymology Volume: 604 Pages: 287-330
  • DOI: 10.1016/bs.mie.2018.02.017
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Discovery of Unprecedented Hydrazine-Forming Machinery in Bacteria (2018)
  • Author(s): Matsuda Kenichi、Tomita Takeo、Shin-ya Kazuo、Wakimoto Toshiyuki、Kuzuyama Tomohisa、Nishiyama Makoto
  • Journal Title: Journal of the American Chemical Society Volume: 140 Issue: 29 Pages: 9083-9086
  • DOI: 10.1021/jacs.8b05354
  • Peer Reviewed
• [Journal Article] Total Synthesis of the Nonribosomal Peptide Surugamide?B and Identification of a New Offloading Cyclase Family (2018)
  • Author(s): Kuranaga Takefumi、Matsuda Kenichi、Sano Ayae、Kobayashi Masakazu、Ninomiya Akihiro、Takada Kentaro、Matsunaga Shigeki、Wakimoto Toshiyuki
  • Journal Title: Angewandte Chemie International Edition Volume: 57 Issue: 30 Pages: 9447-9451
  • DOI: 10.1002/anie.201805541
  • Peer Reviewed
• [Journal Article] Palmitoyl lactic acid induces adipogenesis and a brown fat-like phenotype in 3T3-L1 preadipocytes (2018)
  • Author(s): Uno Yuka、Yamamoto Hirona、Shuto Takatsuki、Sato Yoshinori、Kuranaga Takefumi、Yazawa Kazunaga、Ono Yasuo、Wakimoto Toshiyuki
  • Journal Title: Biochimica et Biophysica Acta (BBA) - Molecular and Cell Biology of Lipids Volume: 1863 Issue: 7 Pages: 772-782
  • DOI: 10.1016/j.bbalip.2018.04.003
  • Peer Reviewed
• [Journal Article] Biosynthetic insights of calyculin- and misakinolide-type compounds in “Candidatus Entotheonella sp.” (2018)
  • Author(s): Uria Agustinus、Piel Joern、Wakimoto Toshiyuki
  • Journal Title: Methods in Enzymology Volume: 604
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Diastereoselective total synthesis and structural confirmation of surugamide F (2018)
  • Author(s): Kuranaga Takefumi、Fukuba Atsuki、Ninomiya Akihiro、Takada Kentaro、Matsunaga Shigeki、Wakimoto Toshiyuki
  • Journal Title: Chemical and Pharmaceutical Bulletin Volume: 66
  • NAID: 130007382323
  • Peer Reviewed
• [Journal Article] Protein phosphatases 1 and 2A and their naturally occurring inhibitors: current topics in smooth muscle physiology and chemical biology (2017)
  • Author(s): Takai Akira、Eto Masumi、Hirano Katsuya、Takeya Kosuke、Wakimoto Toshiyuki、Watanabe Masaru
  • Journal Title: J Physiol Sci Volume: 68 Issue: 1 Pages: 1-17
  • DOI: 10.1007/s12576-017-0556-6
  • NAID: 40021443577
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Total synthesis of theonellapeptolide Id (2017)
  • Author(s): Kuranaga, T., Enomoto, A., Tan, H., Fujita, K., Wakimoto, T.
  • Journal Title: Org. Lett. Volume: 19 Issue: 6 Pages: 1366-1369
  • DOI: 10.1021/acs.orglett.7b00249
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] Toward dark matter of natural products” (2017)
  • Author(s): Wakimoto, T.
  • Journal Title: Chem. Rec. Volume: 17 Issue: 11 Pages: 1124-1134
  • DOI: 10.1002/tcr.201700009
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] Identification of pyridinium with three indole moieties as an antimicrobial agent (2017)
  • Author(s): Okada, M., Sugita, T., Wong, C. P., Wakimoto, T., Abe, I.
  • Journal Title: J. Nat. Prod., Volume: 80 Issue: 4 Pages: 1205-1209
  • DOI: 10.1021/acs.jnatprod.6b01152
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Characterization of giant modular PKSs provides insight into genetic mechanism for structural diversification of aminopolyol polyketides (2017)
  • Author(s): Zhang, L., Hashimoto, T., Qin, B., Hashimoto, J., Kozone, I., Kawahara, T., Okada, M., Awakawa, T., Ito, T., Asakawa, Y., Ueki, M., Takahashi, S., Osada, H., Wakimoto, T., Ikeda, H., Shin-ya, K., Abe, I.
  • Journal Title: Angew. Chem. Int. Ed. Volume: 56 Issue: 7 Pages: 1740-1745
  • DOI: 10.1002/anie.201611371
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Presentation] Investigating spongistatin biosynthesis in complex sponge-bacteria association (2018)
  • Author(s): Agustinus R. Uria, Kenichi Matsuda, Toshiyuki Wakimoto
  • Organizer: 3rd International Symposium on Sponge Microbiology
  • Int'l Joint Research / Invited
• [Presentation] A new off-loading enzyme in non-ribosomal peptide biosynthesis (2018)
  • Author(s): Toshiyuki Wakimoto
  • Organizer: 2nd China-Japan Joint Symposium on Natural Product Biosynthesis
  • Int'l Joint Research / Invited
• [Presentation] Marine Sponge-derived Metagenomes/uncultured Bacteria - a Rich Source of Novel Natural Product Biosynthetic Genes (2018)
  • Author(s): Toshiyuki Wakimoto
  • Organizer: International Seminar on Natural Products Medicines 2018
  • Int'l Joint Research / Invited
• [Presentation] Unusual Processes of PKS and NRPS Assembly Lines in Marine Bacteria (2018)
  • Author(s): Toshiyuki Wakimoto
  • Organizer: 1st German-Japanese Joint Symposium on the Biosynthesis of Natural Products
  • Int'l Joint Research / Invited
• [Presentation] 生合成遺伝子から見出した新規天然物と難培養微生物 (2018)
  • Author(s): 脇本敏幸
  • Organizer: 新化学技術推進協会ライフサイエンス技術部会講演会
  • Invited
• [Presentation] 難培養微生物を起源とする希少医薬品資源の量産 (2018)
  • Author(s): 脇本敏幸
  • Organizer: 生物合成系の再設計による複雑骨格機能分子の革新的創成科学、第２回公開シンポジウム
  • Invited
• [Presentation] メタゲノムマイニングによる海綿由来新規天然物の探索 (2018)
  • Author(s): 脇本敏幸
  • Organizer: 日本薬学会第138会
  • Invited
• [Presentation] メタゲノムマイニングによる海綿由来新規天然物の探索 (2017)
  • Author(s): 脇本敏幸
  • Organizer: 理研シンポジウム、高磁場・高感度NMR利活用促進のための天然物分野における日中連携シンポジウム2017
  • Int'l Joint Research / Invited
• [Presentation] Marine sponge-derived metagenomes/uncultured bacteria - a rich source of novel natural product biosynthetic genes (2017)
  • Author(s): Wakimoto Toshiyuki
  • Organizer: South China Sea Institute of Oceanology, Chinese Academy of Sciences, China
  • Int'l Joint Research / Invited
• [Presentation] Marine sponge-derived metagenomes/uncultured bacteria - a rich source of novel natural product biosynthetic genes (2017)
  • Author(s): Wakimoto Toshiyuki
  • Organizer: Jinan University, College of Pharmacy, China
  • Int'l Joint Research / Invited
• [Presentation] Marine sponge-derived metagenomes/uncultured bacteria - a rich source of novel natural product biosynthetic genes (2017)
  • Author(s): Wakimoto Toshiyuki
  • Organizer: International Conference on Marine Biodiversity and Biotechnology, Jakarta, Indonesia
  • Int'l Joint Research / Invited
• [Presentation] Marine sponge-derived metagenomes/uncultured bacteria - a rich source of novel natural product biosynthetic genes (2017)
  • Author(s): Wakimoto Toshiyuki
  • Organizer: 1st China-Japan Joint Symposium on Natural Product Biosynthesis, Shanghai, China
  • Int'l Joint Research / Invited
• [Presentation] Activated Chemical Defense of Japanese Marine Sponge Discodermia calyx (2017)
  • Author(s): Wakimoto Toshiyuki
  • Organizer: ISCE/APACE, Kyoto, Japan
  • Int'l Joint Research / Invited
• [Presentation] Marine sponge-derived metagenomes/uncultured bacteria - a rich source of novel natural product biosynthetic genes (2017)
  • Author(s): Wakimoto Toshiyuki
  • Organizer: 9th US-Japan Seminar on Natural Product Biosynthesis, UCLA Lake Arrowhead Conference Center, California, USA
  • Int'l Joint Research / Invited
• [Remarks] 北海道大学大学院薬学研究院天然物化学研究室
  • URL: http://www.pharm.hokudai.ac.jp/tennen/index.html
• [Patent(Industrial Property Rights)] ペプチド類の大環状化酵素 (2018)
  • Inventor(s): 脇本敏幸、倉永健史、松田研一
  • Industrial Property Rights Holder: 北海道大学
  • Industrial Property Rights Type: 特許
  • Industrial Property Number: 2019-050797
  • Filing Date: 2018
  • Overseas