Analysis of elementary processes in the photocycle of sodium ion-pumping rhodopsin and an attempt to expand the substrate ion

• Project/Area Number: 17K07326
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Functional biochemistry
• Research Institution: Hokkaido University
• Principal Investigator: Kikukawa Takashi 北海道大学, 先端生命科学研究院, 講師 (20281842)
• Co-Investigator(Kenkyū-buntansha): 宮内 正二 東邦大学, 薬学部, 教授 (30202352)
• Project Period (FY): 2017-04-01 – 2020-03-31
• Project Status: Completed (Fiscal Year 2019)
• Budget Amount: ¥4,680,000 (Direct Cost: ¥3,600,000、Indirect Cost: ¥1,080,000); Fiscal Year 2019: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000); Fiscal Year 2018: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000); Fiscal Year 2017: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
• Keywords: ロドプシン / ナトリウムポンプ / 過渡吸収分光法 / フォトサイクル / 微生物ロドプシン / 膜輸送 / ナノディスク

Outline of Final Research Achievements

The molecular mechanism of Na+-pumping rhodopsin (NaR) was analyzed by employing time-resolved detection techniques and through functional characterization of novel NaR. The results are summarized in two topics. 1) NaR undergoes the photoreaction cycle containing K, L, M, O1, O2, N and NaR´ intermediates, in which Na+ is captured during O1 formation through the hydrated cytoplasmic channel and then released during O2 decay through extracellular channel without further hydration. 2) A novel NaR lacking N-terminal helix can pump not only Na+ but also K+.

Academic Significance and Societal Importance of the Research Achievements

細胞膜には、多くの物質輸送タンパク質が存在し、細胞の恒常性維持に貢献している。本研究で研究対象としたNa+ポンプ型ロドプシン（NaR）は、そのような輸送タンパク質の一つであるが、通常のタンパク質とは異なり、光で瞬間的に活性化できる利点がある。この特徴を生かした解析を行い、Na+輸送時に起こる一部の要素反応を明らかとした。また、新規のNaRの分子特性解析を通して、輸送イオン選択機構の一端を明らかとした。

Research Products

• [Int'l Joint Research] Sogang University(韓国)
• [Journal Article] Functional importance of the oligomer formation of the cyanobacterial H+ pump Gloeobacter rhodopsin (2019)
  • Author(s): Iizuka Azusa、Kajimoto Kousuke、Fujisawa Tomotsumi、Tsukamoto Takashi、Aizawa Tomoyasu、Kamo Naoki、Jung Kwang-Hwan、Unno Masashi、Demura Makoto、Kikukawa Takashi
  • Journal Title: Scientific Reports Volume: 9 Issue: 1 Pages: 10711-10711
  • DOI: 10.1038/s41598-019-47178-5
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Photochemical study of a cyanobacterial chloride-ion pumping rhodopsin (2019)
  • Author(s): Hasemi Takatoshi、Kikukawa Takashi、Watanabe Yumi、Aizawa Tomoyasu、Miyauchi Seiji、Kamo Naoki、Demura Makoto
  • Journal Title: Biochimica et Biophysica Acta (BBA) - Bioenergetics Volume: 1860 Issue: 2 Pages: 136-146
  • DOI: 10.1016/j.bbabio.2018.12.001
  • Peer Reviewed
• [Journal Article] Low-temperature Raman spectroscopy reveals small chromophore distortion in primary photointermediate of proteorhodopsin (2018)
  • Author(s): Fujisawa Tomotsumi、Abe Masahiro、Tamogami Jun、Kikukawa Takashi、Kamo Naoki、Unno Masashi
  • Journal Title: FEBS Letters Volume: 592 Issue: 18 Pages: 3054-3061
  • DOI: 10.1002/1873-3468.13219
  • Peer Reviewed
• [Presentation] Replacements of “H+ donor” residues in the light-driven H+-pump rhodopsins (2019)
  • Author(s): Kikukawa, T., Sasaki, S., Nishiya, K., Tsukamoto, T., Aizawa, T., Demura, M., Tamogami, J.
  • Organizer: 17th International Congress on Photobiology
  • Int'l Joint Research
• [Presentation] Time-resolved detection of Na+ uptake and release reactions of Na+-pumping rhodopsin (2019)
  • Author(s): Murabe, K., Kato, T., Tsukamoto, T., Aizawa, T., Demura, M., Kikukawa, T.
  • Organizer: 9th Asia and Oceania Conference on Photobiology
  • Int'l Joint Research
• [Presentation] Importance of His166 for Cl--pump activity of Mastigocladopsis repens halorhodopsin (2019)
  • Author(s): 岩間健人、渡邊弓、塚本卓、相沢智康、出村誠、菊川峰志
  • Organizer: 第57回日本生物物理学会年会
• [Presentation] イオン選択膜を用いたNa+ポンプ型ロドプシンの機能解析 (2018)
  • Author(s): 村部圭祐, 菊川峰志, 塚本卓, 相沢智康, 出村誠
  • Organizer: 第45回生体分子科学討論会
• [Presentation] Photochemical analysis of sodium ion-pumping rhodopsin from Indibacter alkaliphilus (2018)
  • Author(s): 加藤 朝也, 村部 圭祐, 菊川 峰志, 塚本 卓, 相沢 智康, 出村 誠
  • Organizer: 第56回日本生物物理学会年会
• [Presentation] Functional importance of trimer formation of light-driven H+ pump Gloeobacter rhodopsin (2018)
  • Author(s): Kikukawa, T.
  • Organizer: 18th International Conference on Retinal Proteins
  • Int'l Joint Research
• [Presentation] Cyanobacterial rhodopsin having TSD motif (2017)
  • Author(s): Kikukawa, T.
  • Organizer: 8th Asia and Oceania Conference on Photobiology
  • Int'l Joint Research / Invited
• [Presentation] Photochemistry of cyanobacterial chloride ion-pumping rhodopsin (2017)
  • Author(s): Kikukawa, T., Hasemi, T., Aizawa, T., Kamo, N., Demura, M.
  • Organizer: 19th International Union for Pure and Applied Biophysics Congress and 11th European Biophysics Congress
  • Int'l Joint Research
• [Presentation] Functional analyses of Na+-pumping rhodopsin focusing on acidic residues on the extracellular surface (2017)
  • Author(s): Okamura, A., Kikukawa, T., Tsukamoto, T., Aizawa, T., Kamo, N., Demura, M.
  • Organizer: 第５５回日本生物物理学会年会
• [Presentation] Analysis of Na+ transfer reactions of Na+-pumping rhodopsin (2017)
  • Author(s): Murabe, K., Kikukawa, T., Tsukamoto, T., Aizawa, T., Kamo, N., Demura, M.
  • Organizer: 第５５回日本生物物理学会年会
• [Presentation] Functional importance of trimer formation of light-driven H+ pump Gloeobacter rhodopsin (2017)
  • Author(s): Iizuka, A., Kikukawa, T., Kajimoto, K., Fujisawa, T., Tsukamoto, T., Aizawa, T., Kamo, N., Unno, M., Demura, M.
  • Organizer: 第５５回日本生物物理学会年会
• [Presentation] Functional analysis of T126E mutant of Natronomonas pharaonis halorhodopsin (2017)
  • Author(s): Abe, S., Kikukawa, T., Tsukamoto, T., Aizawa, T., Kamo, N., Demura, M.
  • Organizer: 第５５回日本生物物理学会年会