フロリゲン複合体の立体構造解析と低分子化合物による花成制御

• Project Area: Determining the principles of the birth of new plant species: molecular elucidation of the lock-and-key systems in sexual reproduction
• Project/Area Number: 17H05836
• Research Category: Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area)
• Allocation Type: Single-year Grants
• Review Section: Biological Sciences
• Research Institution: Yokohama National University
• Principal Investigator: 児嶋 長次郎 横浜国立大学, 大学院工学研究院, 教授 (50333563)
• Project Period (FY): 2017-04-01 – 2019-03-31
• Project Status: Completed (Fiscal Year 2018)
• Budget Amount: ¥11,310,000 (Direct Cost: ¥8,700,000、Indirect Cost: ¥2,610,000); Fiscal Year 2018: ¥5,590,000 (Direct Cost: ¥4,300,000、Indirect Cost: ¥1,290,000); Fiscal Year 2017: ¥5,720,000 (Direct Cost: ¥4,400,000、Indirect Cost: ¥1,320,000)
• Keywords: NMR / 蛋白質 / 立体構造

Outline of Annual Research Achievements

本研究では、花成ホルモン・受容体・転写因子・DNA4者複合体の立体構造解析と、花成の制御が可能な低分子化合物の創製によって、植物の生産性や生体機能の 精密制御の基盤とすることを目的としている。平成29年度から継続して、(1)全自動NMR構造解析システムMagROの開発、(2)花成ホルモン受容体をターゲットとしたスクリーニングと化合物最適化、を進めるとともに、独自のアミノ酸選択標識技術などと組み合わせ、平成30年度から新たに、(3)複合体構成蛋白質単体の立体構造決定、(4)花成ホルモン受容体複合体と薬剤複合体の構造解析、に取り組んだ。本年度の具体的な研究成果は以下の通り。
(1)昨年度開発した全自動NMR構造解析システムについては、様々な蛋白質への適用が可能であることを確認した。また、花成ホルモンと受容体に適用した。(2)受容体-転写因子相互作用を標的とし、4万化合物から花成制御化合物を得ることに成功した。しかし得られた化合物の類縁化合物では溶解度を向上させることができなかったため、標的とすべき蛋白質間相互作用を変えて再度スクリーニングを行った。(3)(4)立体構造解析では、花成ホルモン・受容体・転写因子・DNAの4者複合体をハイブリッド法で立体構造解析するための基盤として、複合体の構成蛋白質個々に単体での立体構造決定と複合体の安定化を進めた。手法としてX線とNMRの両方を用い、花成ホルモンと受容体・転写因子複合体の構造解析を進め、受容体・転写因子複合体については立体構造決定に成功した。低分子化合物の創製では、19F-NMRを活用した蛋白質薬剤構造構築法を新たに開発した。また、花成ホルモンと薬剤との相互作用解析を進めた。

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] Academy of Sciences(チェコ)
• [Int'l Joint Research] Marquette University(米国)
• [Int'l Joint Research] Universite de Sherbrooke(Canada)
• [Journal Article] The Helix Rearrangement in the Periplasmic Domain of the Flagellar Stator B Subunit Activates Peptidoglycan Binding and Ion Influx (2018)
  • Author(s): Kojima Seiji, Takao Masato, Almira Gaby, Kawahara Ikumi, Sakuma Mayuko, Homma Michio, Kojima Chojiro, Imada Katsumi.
  • Journal Title: Structure Volume: 26 Issue: 4 Pages: 590-598
  • DOI: 10.1016/j.str.2018.02.016
  • Peer Reviewed
• [Journal Article] Amino Acid Selective 13C Labeling and 13C Scrambling Profile Analysis of Protein α and Side-Chain Carbons in Escherichia coli Utilized for Protein Nuclear Magnetic Resonance (2018)
  • Author(s): Sugiki Toshihiko、Furuita Kyoko、Fujiwara Toshimichi、Kojima Chojiro
  • Journal Title: Biochemistry Volume: 57 Issue: 26 Pages: 3576-3589
  • DOI: 10.1021/acs.biochem.8b00182
  • Peer Reviewed
• [Journal Article] Phosphoinositide binding by the PH domain in ceramide transfer protein (CERT) is inhibited by hyperphosphorylation of an adjacent serine-repeat motif (2018)
  • Author(s): Sugiki Toshihiko、Egawa Daichi、Kumagai Keigo、Kojima Chojiro、Fujiwara Toshimichi、Takeuchi Koh、Shimada Ichio、Hanada Kentaro、Takahashi Hideo
  • Journal Title: Journal of Biological Chemistry Volume: 293 Issue: 28 Pages: 11206-11217
  • DOI: 10.1074/jbc.ra118.002465
  • Peer Reviewed / Open Access
• [Journal Article] NMR spectroscopic characterization of a model RNA duplex reflecting the core sequence of hammerhead ribozymes (2018)
  • Author(s): Hattori Yoshikazu、Yamanaka Daichi、Morioka Saori、Yamaguchi Taishi、Tomonari Honoka、Kojima Chojiro、Tanaka Yoshiyuki
  • Journal Title: Nucleosides, Nucleotides and Nucleic Acids Volume: 37 Issue: 7 Pages: 383-396
  • DOI: 10.1080/15257770.2018.1473877
  • Peer Reviewed
• [Journal Article] Noise peak filtering in multi-dimensional NMR spectra using convolutional neural networks (2018)
  • Author(s): Kobayashi, N., Hattori, Y., Nagata, T., Shinya, S., Guentert, P., Kojima, C., Fujiwara, T.
  • Journal Title: Bioinformatics Volume: 34 Issue: 24 Pages: 4300-4301
  • DOI: 10.1093/bioinformatics/bty581
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] TFL1-Like Proteins in Rice Antagonize Rice FT-Like Protein in Inflorescence Development by Competition for Complex Formation with 14-3-3 and FD. (2018)
  • Author(s): Kaneko-Suzuki M, Kurihara-Ishikawa R, Okushita-Terakawa C, Kojima C, Nagano-Fujiwara M, Ohki I, Tsuji H, Shimamoto K, Taoka KI.
  • Journal Title: Plant Cell Physiol. Volume: 59 Issue: 3 Pages: 458-468
  • DOI: 10.1093/pcp/pcy021
  • Peer Reviewed / Open Access
• [Journal Article] Current NMR Techniques for Structure-Based Drug Discovery (2018)
  • Author(s): Sugiki Toshihiko、Furuita Kyoko、Fujiwara Toshimichi、Kojima Chojiro
  • Journal Title: Molecules Volume: 23 Issue: 1 Pages: 148-148
  • DOI: 10.3390/molecules23010148
  • Peer Reviewed / Open Access
• [Journal Article] Production of Single-Chain Fv Antibodies Specific for GA-Pyridine, an Advanced Glycation End-Product (AGE), with Reduced Inter-Domain Motion (2017)
  • Author(s): Fukuda N、Noi K、Weng L、Kobashigawa Y、Miyazaki H、Wakeyama Y、Takaki M、Nakahara Y、Tatsuno Y、Uchida-Kamekura M、Suwa Y、Sato T、Ichikawa-Tomikawa N、Nomizu M、Fujiwara Y、Ohsaka F、Saitoh T、Maenaka K、Kumeta H、Shinya S、Kojima C、Ogura T、Morioka H
  • Journal Title: Molecules Volume: 22 Issue: 10 Pages: 1695-1695
  • DOI: 10.3390/molecules22101695
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Structural and Functional Analysis of the C-Terminal Region of FliG, an Essential Motor Component of Vibrio Na + -Driven Flagella (2017)
  • Author(s): Miyanoiri Yohei、Hijikata Atsushi、Nishino Yuuki、Gohara Mizuki、Onoue Yasuhiro、Kojima Seiji、Kojima Chojiro、Shirai Tsuyoshi、Kainosho Masatsune、Homma Michio
  • Journal Title: Structure Volume: 25 Issue: 10 Pages: 1540-1548
  • DOI: 10.1016/j.str.2017.08.010
  • NAID: 120006375873
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Protein 19F-labeling using transglutaminase for the NMR study of intermolecular interactions (2017)
  • Author(s): Hattori Yoshikazu、Heidenreich David、Ono Yuki、Sugiki Toshihiko、Yokoyama Kei-ichi、Suzuki Ei-ichiro、Fujiwara Toshimichi、Kojima Chojiro
  • Journal Title: Journal of Biomolecular NMR Volume: 68 Issue: 4 Pages: 271-279
  • DOI: 10.1007/s10858-017-0125-6
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] The mechanism of the glycosylase reaction with hOGG1 base-excision repair enzyme: Concerted effect of Lys249 and Asp268 during excision of 8-oxoguanine. (2017)
  • Author(s): Jakub Sebera, Yoshikazu Hattori, Daichi Sato, David Reha, Radim Nencka, Takashi Kohno, Chojiro Kojima, Yoshiyuki Tanaka, and Vladimir Sychrovsky
  • Journal Title: Nucleic Acids Research Volume: 45 Issue: 9 Pages: 5231-5242
  • DOI: 10.1093/nar/gkx157
  • Peer Reviewed / Open Access / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] NMR line shape analysis of a multi-state ligand binding mechanism in chitosanase (2017)
  • Author(s): Shinya Shoko、Ghinet Mariana G.、Brzezinski Ryszard、Furuita Kyoko、Kojima Chojiro、Shah Sneha、Kovrigin Evgenii L.、Fukamizo Tamo
  • Journal Title: Journal of Biomolecular NMR Volume: 67 Issue: 4 Pages: 309-319
  • DOI: 10.1007/s10858-017-0109-6
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Cold-Shock Expression System in E. coli for Protein NMR Studies (2017)
  • Author(s): Sugiki Toshihiko、Fujiwara Toshimichi、Kojima Chojiro
  • Journal Title: Methods Mol Biol Volume: 1586 Pages: 345-357
  • DOI: 10.1007/978-1-4939-6887-9_23
  • ISBN: 9781493968855, 9781493968879
  • Peer Reviewed
• [Presentation] 創薬 NMR 技術の開発とその植物生理学分野への応用 (2018)
  • Author(s): 児嶋長次郎
  • Organizer: 日本蛋白質科学会第18回年会
  • Invited
• [Presentation] NMR tools developed for protein-drug and protein-protein interaction studies (2018)
  • Author(s): Chojiro Kojima
  • Organizer: The 6th International Symposium on Drug Discovery and Design by NMR
  • Int'l Joint Research / Invited
• [Presentation] 植物構造生物学研究と創薬 (2018)
  • Author(s): 児嶋長次郎
  • Organizer: 植物科学シンポジウム 2018
  • Invited
• [Remarks] Kojima Lab top page
  • URL: http://nmrlab.ynu.ac.jp/labotop.php