Unique hydrogen-bonding environments in photosensor proteins

• Project/Area Number: 16K07318
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Biophysics
• Research Institution: Toho University
• Principal Investigator: IWATA Tatsuya 東邦大学, 薬学部, 准教授 (20569917)
• Project Period (FY): 2016-04-01 – 2019-03-31
• Project Status: Completed (Fiscal Year 2018)
• Budget Amount: ¥4,940,000 (Direct Cost: ¥3,800,000、Indirect Cost: ¥1,140,000); Fiscal Year 2018: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000); Fiscal Year 2017: ¥1,690,000 (Direct Cost: ¥1,300,000、Indirect Cost: ¥390,000); Fiscal Year 2016: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
• Keywords: フラビン / 光センサータンパク質 / フーリエ変換赤外分光法 / 安定同位体標識 / 質量分析法 / 光受容体 / フーリエ変換赤外（FTIR）分光法 / BLUFドメイン / LOVドメイン / 水素結合

Outline of Final Research Achievements

Formation of specific hydrogen bonds was detected by infrared spectroscopy on blue-light receptor domains, LOV and BLUF, that binds flavin as chromophore.
I found the NH group of flavin forms quite strong hydrogen bond with side chains of apoprotein in LOV domain. Flavin must bind into the LOV apoprotein by such a strong interaction. For the BLUF domain, we have found quite strong hydrogen bond of the OH group of Tyr side chain, and the partner was a nitrogen atom of enol form of Gln side chain.

Academic Significance and Societal Importance of the Research Achievements

本研究では、赤外分光法を用いてタンパク質中の強い水素結合を形成した化学結合の検出に成功しました。水素結合はタンパク質の構造形成や基質認識や酵素反応に重要な役割を果たしていると考えられますが、それを検出できる系というのは限られています。本研究で発見したことは、光センサータンパク質の光反応を測定することによって、LOVドメインではタンパク質と補酵素の結合で恒常的に強い水素結合を形成すること、またBLUFドメインでは光反応中間体で強い水素結合を形成することを見出しました。今後、適切にな測定系の開発により、多くのタンパク質の構造形成や反応に重要な水素結合の検出への足がかりになるものと考えられます。

Research Products

• [Int'l Joint Research] スクリプス研究所(米国)
• [Journal Article] Hydrogen Bonding Environments in the Photocycle Process around the Flavin Chromophore of the AppA-BLUF domain (2018)
  • Author(s): Iwata Tatsuya、Nagai Takashi、Ito Shota、Osoegawa Shinsuke、Iseki Mineo、Watanabe Masakatsu、Unno Masashi、Kitagawa Shinya、Kandori Hideki
  • Journal Title: Journal of the American Chemical Society Volume: 140 Issue: 38 Pages: 11982-11991
  • DOI: 10.1021/jacs.8b05123
  • Peer Reviewed / Open Access
• [Journal Article] Hydrogen Bonding Environment of the N3?H Group of Flavin Mononucleotide in the Light Oxygen Voltage Domains of Phototropins (2017)
  • Author(s): Iwata Tatsuya、Nozaki Dai、Yamamoto Atsushi、Koyama Takayuki、Nishina Yasuzo、Shiga Kiyoshi、Tokutomi Satoru、Unno Masashi、Kandori Hideki
  • Journal Title: Biochemistry Volume: 56 Issue: 24 Pages: 3099-3108
  • DOI: 10.1021/acs.biochem.7b00057
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Functional Conversion of CPD and (6-4) Photolyases by Mutation (2016)
  • Author(s): Daichi Yamada, Hisham M. Dokainish, Tatsuya Iwata, Junpei Yamamoto, Tomoko Ishikawa, Takeshi Todo, Shigenori Iwai, Elizabeth D. Getzoff, Akio Kitao, and Hideki Kandori
  • Journal Title: Biochemistry Volume: 55 Issue: 30 Pages: 4173-4183
  • DOI: 10.1021/acs.biochem.6b00361
  • Peer Reviewed / Open Access / Int'l Joint Research / Acknowledgement Compliant
• [Presentation] PHR/CRY ファミリーにおける FAD 酸化還元状態の制御メカニズムの解析 (2017)
  • Author(s): ○酒井結衣、山田大智、伊藤 奨太、岩田達也、神取秀樹
  • Organizer: 日本生物物理学会中部支部講演会
  • Place of Presentation: 名古屋大学東山キャンパス（愛知県・名古屋市）
  • Year and Date: 2017-03-06
• [Presentation] Artificial photosynthesis based on the engineered flavoprotein LOV (2016)
  • Author(s): ○N. Ueda, Y. Ono, T. Iwata;M. Iwaki,H. Kandori
  • Organizer: The 54th Annual Meeting of the Biophysical Society of Japan
  • Place of Presentation: つくば国際会議場（茨城県・つくば市）
  • Year and Date: 2016-11-25
• [Presentation] FTIR spectroscopic analysis of a DNAzyme possessing DNA photorepair activity (2016)
  • Author(s): ○Y. Kurahashi;I M. M. Wijaya, T. Iwata;H. Kandori
  • Organizer: The 54th Annual Meeting of the Biophysical Society of Japan
  • Place of Presentation: つくば国際会議場（茨城県・つくば市）
  • Year and Date: 2016-11-25
• [Presentation] Analysis of a hydrogen bonding network of the BLUF domain using isotope-labeled samples (2016)
  • Author(s): ○T. Nagai, T. Iwata;S. Ito, M. Iseki, M. Watanabe, M. Unno, S. Kitagawa,H. Kandori
  • Organizer: The 54th Annual Meeting of the Biophysical Society of Japan
  • Place of Presentation: つくば国際会議場（茨城県・つくば市）
  • Year and Date: 2016-11-25
• [Presentation] Comparative FTIR study of photoactivation and photorepair of T(6-4)T and T(6-4)C photoproducts by Xenopus (6-4) photolyase (2016)
  • Author(s): ○M. Kumagai, D. Yamada, T. Iwata, J. Yamamoto, E. D. Getzoff, S. Iwai;H. Kandori
  • Organizer: The 54th Annual Meeting of the Biophysical Society of Japan
  • Place of Presentation: つくば国際会議場（茨城県・つくば市）
  • Year and Date: 2016-11-25
• [Presentation] FTIR study of sodium binding and photoreaction in KR2 (2016)
  • Author(s): ○S. Ito;R. Abe-Yoshizumi;S. Sugita,K. Inoue;T. Iwata;M. Iwaki;H. Kandori
  • Organizer: The 54th Annual Meeting of the Biophysical Society of Japan
  • Place of Presentation: つくば国際会議場（茨城県・つくば市）
  • Year and Date: 2016-11-25
• [Presentation] How are flavoproteins "soft" for exhibiting intended functions? (2016)
  • Author(s): T. Iwata
  • Organizer: The 54th Annual Meeting of the Biophysical Society of Japan
  • Place of Presentation: つくば国際会議場（茨城県・つくば市）
  • Year and Date: 2016-11-25
  • Invited
• [Presentation] Photochemical properties of DNA-bound flavin (2016)
  • Author(s): ○T. Iwata, M. Hayakawa,H. Kandori
  • Organizer: The 43rd international symposium on Nucleic Acids chemistry
  • Place of Presentation: 熊本大学（熊本県・熊本市）
  • Year and Date: 2016-09-27
• [Presentation] Structural analysis of DNAzyme possessing DNA photolyase activity by FTIR spectroscopy (2016)
  • Author(s): ○Y. Kurahashi;I M. M. Wijaya,T. Iwata;H. Kandori
  • Organizer: The 43rd international symposium on Nucleic Acids chemistry
  • Place of Presentation: 熊本大学（熊本県・熊本市）
  • Year and Date: 2016-09-27
• [Presentation] Infrared spectrum analysis of a DNAzyme that functions as photorepair of UV-induced DNA (2016)
  • Author(s): ○Y. Kurahashi;I M. M. Wijaya, T. Iwata, H. Kandori
  • Organizer: The 4th Awaji International Workshop on "Electron Spin Science & Technology: Biological and Materials Science Oriented Applications
  • Place of Presentation: 淡路夢舞台（兵庫県・洲本市）
  • Year and Date: 2016-06-19
  • Int'l Joint Research
• [Presentation] How can a neutral semiquinone radical of flavin be stably formed in DNA photolyase? (2016)
  • Author(s): T. Iwata
  • Organizer: The 4th Awaji International Workshop on "Electron Spin Science & Technology: Biological and Materials Science Oriented Applications
  • Place of Presentation: 淡路夢舞台（兵庫県・洲本市）
  • Year and Date: 2016-06-19
  • Int'l Joint Research / Invited
• [Presentation] Comparison of photoactivation and photorepair of T(6-4)T and T(6-4)C photoproducts by Xenopus (6-4) photolyase on FTIR study (2016)
  • Author(s): ○M. Kumagai, D. Yamada,T. Iwata, J. Yamamoto, E. D. Getzoff, S. Iwai,H. Kandori
  • Organizer: American Society for Photobiology Conference
  • Place of Presentation: タンパ（アメリカ合衆国・フロリダ州）
  • Year and Date: 2016-05-21
  • Int'l Joint Research
• [Presentation] Artificial photosynthesis with a single redox centre based on the engineered flavoprotein LOV (2016)
  • Author(s): ○N. Ueda, Y. Ono;T. Iwata;M. Iwaki;H. Kandori
  • Organizer: American Society for Photobiology Conference
  • Place of Presentation: タンパ（アメリカ合衆国・フロリダ州）
  • Year and Date: 2016-05-21
  • Int'l Joint Research