Development of fully-automated system for NMR structural analysis of higher molecular weight proteins

• Project/Area Number: 17H03641
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Structural biochemistry
• Research Institution: Yokohama National University
• Principal Investigator: KOJIMA Chojiro 横浜国立大学, 大学院工学研究院, 教授 (50333563)
• Project Period (FY): 2017-04-01 – 2020-03-31
• Project Status: Completed (Fiscal Year 2019)
• Budget Amount: ¥17,420,000 (Direct Cost: ¥13,400,000、Indirect Cost: ¥4,020,000); Fiscal Year 2019: ¥5,200,000 (Direct Cost: ¥4,000,000、Indirect Cost: ¥1,200,000); Fiscal Year 2018: ¥5,850,000 (Direct Cost: ¥4,500,000、Indirect Cost: ¥1,350,000); Fiscal Year 2017: ¥6,370,000 (Direct Cost: ¥4,900,000、Indirect Cost: ¥1,470,000)
• Keywords: NMR / 蛋白質 / 立体構造

Outline of Final Research Achievements

In this research, we have developed a system for determining the NMR structure of higher molecular weight proteins by making use of 950 MHz NMR equipment and fully automated structure calculation software. Specifically, the utilizitation of fully automated NMR structural determination system MagRO, amino acid selective labeling technology, and methyl group-selective deuterium labeling has enabled the structural analysis of protein-protein and protein-drug complexes.

Academic Significance and Societal Importance of the Research Achievements

NMRによる蛋白質の立体構造決定は専門性が高く、今まで専門家以外の研究者が利用することが難しかった。本研究では、大阪大学蛋白質研究所で整備されている世界最高感度のNMR装置、小林直宏博士（現理化学研究所）が開発した全自動NMR自動構造計算ソフト、独自開発の安定同位体標識技術などを用いることで、高分子量蛋白質にNMR構造解析が適用可能であることを示した。今後は蛋白質のNMR構造解析の汎用化が進むと考えられる。

Research Products

• [Int'l Joint Research] Rutgers University(米国)
• [Int'l Joint Research] Goethe University Frankfurt(ドイツ)
• [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(カナダ)
• [Journal Article] Crystal contact-free conformation of an intrinsically flexible loop in protein crystal: Tim21 as the case study (2020)
  • Author(s): Bala Siqin、Shinya Shoko、Srivastava Arpita、Ishikawa Marie、Shimada Atsushi、Kobayashi Naohiro、Kojima Chojiro、Tama Florence、Miyashita Osamu、Kohda Daisuke
  • Journal Title: Biochimica et Biophysica Acta (BBA) - General Subjects Volume: 1864 Issue: 2 Pages: 129418-129418
  • DOI: 10.1016/j.bbagen.2019.129418
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] In-cell NMR as a sensitive tool to monitor physiological condition of Escherichia coli (2020)
  • Author(s): Sugiki Toshihiko、Yamaguchi Yoshihiro、Fujiwara Toshimichi、Inouye Masayori、Ito Yutaka、Kojima Chojiro
  • Journal Title: Scientific Reports Volume: 10 Issue: 1 Pages: 2466-2466
  • DOI: 10.1038/s41598-020-59076-2
  • NAID: 120006952208
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Structural instability of lamin A tail domain modulates its assembly and higher order function in Emery-Dreifuss muscular dystrophy (2019)
  • Author(s): Muneyo Mio, Toshihiko Sugiki, Chie Matsuda, Hiroaki Mitsuhashi, Chojiro Kojima, Siu Yuen Chan, Yukiko K. Hayashi, Kazuhiro Mio
  • Journal Title: Biochemical and Biophysical Research Communications Volume: 512 Issue: 1 Pages: 22-28
  • DOI: 10.1016/j.bbrc.2019.02.138
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Structural characterization of the N-terminal kinase-interacting domain of an Hsp90-cochaperone Cdc37 by CD and solution NMR spectroscopy. (2019)
  • Author(s): Ihama, F., Yamamoto, M., Kojima, C., Fujiwara, T., Matsuzaki, K., Miyata, Y., Hoshino, M.
  • Journal Title: Biochim. Biophys. Acta-Proteins Proteom. Volume: 1867 Issue: 9 Pages: 813-820
  • DOI: 10.1016/j.bbapap.2019.06.007
  • 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] 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] 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] 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] 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] 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] 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] 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] 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] 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
• [Presentation] 19F-NMR for Fragment-Based Drug Discovery (2019)
  • Author(s): Chojiro Kojima
  • Organizer: 8th Asia-Pacific NMR Symposium 2019
  • Int'l Joint Research / Invited
• [Presentation] NMRで分子集団の構造を捉える (2019)
  • Author(s): 児嶋長次郎
  • Organizer: 第20回若手NMR研究会
  • Invited
• [Presentation] 19F-NMR for Drug Discovery (2019)
  • Author(s): Chojiro Kojima
  • Organizer: Chem-Bio Informatics Society (CBI) Annual Meeting 2019
  • Invited
• [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
• [Presentation] Fully Automated NMR Structure Determination of Protein (2017)
  • Author(s): Chojiro Kojima
  • Organizer: 2017 Taiwan-Japan Biomedical Symposium on Magnetic Resonance
  • Int'l Joint Research / Invited
• [Remarks] Kojima Lab top page
  • URL: http://nmrlab.ynu.ac.jp/labotop.php