X-ray crystallographic study of squid rhodopsin

• Project/Area Number: 19770129
• Research Category: Grant-in-Aid for Young Scientists (B)
• Allocation Type: Single-year Grants
• Research Field: Biophysics
• Research Institution: Nagoya University
• Principal Investigator: MURAKAMI Midori Nagoya University, 大学院・理学研究科, 助教 (20324387)
• Project Period (FY): 2007 – 2008
• Project Status: Completed (Fiscal Year 2008)
• Budget Amount: ¥3,530,000 (Direct Cost: ¥3,200,000、Indirect Cost: ¥330,000); Fiscal Year 2008: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2007: ¥2,100,000 (Direct Cost: ¥2,100,000)
• Keywords: ロドプシン / X線結晶構造解析 / GPCR

Research Abstract

視物質ロドプシンは網膜視細胞に存在する光受容膜蛋白質である。無脊椎動物のロドプシンは、脊椎動物のロドプシンとは対照的に光活性型アシッドメタ中間体が安定であり、また、生体内に普遍的に存在するGq型G蛋白質と共役するという特徴をもつ。研究代表者らはスルメイカのロドプシンについてX線結晶構造解析を行い、2007年度までに基底状態の立体構造を2.5Å分解能で決定し、無脊椎動物型ロドプシンの機能を裏付ける以下のような特徴的な構造を解明した。1)GPCRの膜貫通領域はほぼ保存されている、2)細胞質側のG蛋白質共役部位が堅いドメイン構造である、3)9個の水分子クラスタが活性部位とG蛋白質共役部位を結ぶ水素結合網を形成し、細胞内シグナル伝達に寄与している、4)発色団レチナールのプロトン化シッフ塩基とその対イオンGlu180は5Å離れている、5)GPCRの生理的な二量体がへリックス4と5を接合面として会合している、6)細胞外側領域どうしの塩橋による相互作用が無脊椎動物の目が偏光を検出するのに寄与している。2008年度は光反応初期中間体であるバソロドプシンの結晶構造解析を目指し、イカロドプシン結晶中でバソ反応中間体を捕捉する条件検討を行った。すなわち、液体窒素温度に冷却した結晶に対し、青色光照射によりバソ反応中間体(全トランス型)と、橙色光照射によりロドプシン(11-cis 型)とイソロドプシン(9-cis型)の混合状態とを可逆的に生成することを見出した。バソ中間体の生成率が高めた結晶を用いて回折データセットを収集し、構造解析に着手した。

Research Products

• [Journal Article] Functionally important structural elements of the cyanobacterial clock-related protein Pex (2009)
  • Author(s): S. Kurosawa, R. Murakami, K. Onai, M. Morishita, D. Hasegawa, R. Iwase, T. Uzumaki, F. Hayashi, T. Kitajima-Ihara, S. Sakata, M. Murakami, T. Kouyama and M. Ishiura
  • Journal Title: Genes Cells 14 Pages: 1-16
  • Peer Reviewed
• [Journal Article] イカロドプシンの結晶構造と無脊椎動物の視覚 (2009)
  • Author(s): 村上緑
  • Journal Title: 生物物理 49 Pages: 15-16
  • NAID: 110007044738
  • Peer Reviewed
• [Journal Article] Functionally important structural elements of the cyanobacterial clock-related protein Pex (2009)
  • Author(s): S. Kurosawa, R. Murakami, K. Onei, M. Morishita, D. Hasegalira, R. Iwase, T. Uzumaki, F. Hayashi, T. Kitajima-Ihara, S. Sakata, M. Murakami, T. Kouyama and M. Ishiura
  • Journal Title: Genes Cells 14 Pages: 1-16
  • Peer Reviewed
• [Journal Article] Effect of xenon binding to a hydrophobic cavity on the proton pumping cycle in bacteriorhodopsin (2008)
  • Author(s): N. Hayakawa, T. Kasahara, D. Hasegawa, K. Yoshimura, M. Murakami and T. Kouyama
  • Journal Title: J. Mol. Biol. 384 Pages: 812-823
  • Peer Reviewed
• [Journal Article] スルメイカロドプシンのX線結晶構造解析 (2008)
  • Author(s): 村上緑, 神山勉
  • Journal Title: 実験医学 26 Pages: 2237-2240
  • Peer Reviewed
• [Journal Article] Crystal structure of squid rhodopsin (2008)
  • Author(s): M. Murakami and T. Kouyama
  • Journal Title: Nature 453 Pages: 363-367
  • Peer Reviewed
• [Journal Article] Crystal structure of squid rhodopsin (2008)
  • Author(s): M. Murakami and T. Kouyaina
  • Journal Title: Nature 453 Pages: 363-367
  • Peer Reviewed
• [Journal Article] スルメイカロドプシンのX線結晶構造解析 (2008)
  • Author(s): 村上緑、神山勉
  • Journal Title: 実験医学 26 Pages: 2237-2240
  • Peer Reviewed
• [Journal Article] Crystallization and crystal properties of squid rhodopsin (2007)
  • Author(s): M. Murakami, R. Kitahara, T. Gotoh and T. Kouyama
  • Journal Title: Acta Cryst F63 Pages: 475-479
  • Peer Reviewed
• [Journal Article] Crystallization and crystal properties of squid rhodopsin (2007)
  • Author(s): M. Murakami, R. Kitahara, T. Gotoh and T. Kouyama
  • Journal Title: Acta Cryst. F63 Pages: 475-279
  • Peer Reviewed
• [Presentation] イカロドプシン光反応初期中間体の構造解析 (2009)
  • Author(s): 村上緑、神山勉
  • Organizer: 第4回日本生物物理学会中部支部討論会
  • Place of Presentation: 名古屋
  • Year and Date: 2009-03-31
• [Presentation] イカロドプシン光異性化反応の構造解析 (2009)
  • Author(s): 村上緑
  • Organizer: シンポジウム : 名大の生物物理の生誕50周年
  • Place of Presentation: 名古屋
  • Year and Date: 2009-03-26
• [Presentation] X-ray structure of squid rhodopsin (2008)
  • Author(s): M. Murakami
  • Organizer: International symposium on membrane proteins and high resolution X-ray structural analysis
  • Place of Presentation: Hyogo
  • Year and Date: 2008-09-01
• [Presentation] Crystal Structure of Squid Rhodopsin (2008)
  • Author(s): M. Murakami
  • Organizer: Gordon Research Conference on "LIGAND RECOGNITION&MOLECULAR GATING"
  • Place of Presentation: Ventura, CA, USA
  • Year and Date: 2008-03-06
• [Presentation] Crystal Structure of Squid Rhodopsin at 2.5Å resolution (2008)
  • Author(s): T. Kouyama and M. Murakami
  • Organizer: GPCR symposium -Frontier on Integral Membrane Proteins in Crystal Structure Determination-
  • Place of Presentation: Hyogo
  • Year and Date: 2008-02-26
• [Presentation] バクテリオロドプシンのアルカリpHでの光誘起構造変化 (2008)
  • Author(s): 山本昌孝、村上緑、神山勉
  • Organizer: 日本生物物理学会第46回年会
  • Place of Presentation: 福岡
• [Presentation] イカロドプシンのX線結晶構造解析 (2008)
  • Author(s): 村上緑、神山勉
  • Organizer: 日本生物物理学会第46回年会
  • Place of Presentation: 福岡
• [Presentation] Crystallization of visual pigments and archaeal rhodopsins (2008)
  • Author(s): T. Kouyama, M. Murakami, K. Yoshimura and M. Yamamoto
  • Organizer: XXI Congress and General Assembly of the International Union of Crystallography
  • Place of Presentation: Osaka
• [Presentation] Dimeric structure of squid rhodopsin (2008)
  • Author(s): M. Murakami and T. Kouyama
  • Organizer: 13th International conference of retinal proteins
  • Place of Presentation: Barcelona
• [Presentation] Protein-lipid interactions in 3D crystals of squid rhodopsin and archaeal rhodopsins (2008)
  • Author(s): T. Kouyama, M. Murakami and K. Yoshimura
  • Organizer: 13th International conference of retinal proteins
  • Place of Presentation: Barcelona
• [Presentation] スルメイカロドプシンのX線結晶構造解析 (2008)
  • Author(s): 村上緑、神山勉
  • Organizer: 第8回日本蛋白質科学会年会
  • Place of Presentation: 東京
• [Presentation] Crystal Structure of Squid Rhodopsin (2008)
  • Author(s): M. Murakami
  • Organizer: Gordon Research Conference on "LIGAND RECOGNITION & MOLECULAR GATING"
  • Place of Presentation: Ventura, CA
• [Presentation] Protein-lipid interactions in 3D crystals of squid rhodopsin and archaeal rhodopsins (2008)
  • Author(s): T. Kouyama, M. Murakami and K. Yoshimura
  • Organizer: 13 th International conference of retinal proteins
  • Place of Presentation: Barcelona
• [Presentation] Dimeric structure of squid rhodopsin (2008)
  • Author(s): M. Murakami and T. Kouyama
  • Organizer: 13 th International conference of retinal proteins
  • Place of Presentation: Barcelona
• [Presentation] Crystal Structure of Squid Rhodopsin (2007)
  • Author(s): M. Murakami and T. Kouyama
  • Organizer: International Symposium on RETINAL PROTEINS: EXPERIMENTS AND THEORY
  • Place of Presentation: Bremen, Germany
  • Year and Date: 2007-09-24
• [Presentation] X-ray crystallographic study of squid rhodopsin (2007)
  • Author(s): M. Murakami and T. Kouyama
  • Organizer: 2nd International Symposium on Diffraction Structural Biology 2007
  • Place of Presentation: 東京
  • Year and Date: 2007-09-10
• [Presentation] バクテリオロドプシンの疎水キャビティに結合する希ガスとその効果 (2007)
  • Author(s): 早川直紀、村上緑、神山勉
  • Organizer: 日本生物物理学会第45回年会
  • Place of Presentation: 横浜
• [Presentation] バクテリオロドプシン3次元結晶のM中間体寿命に及ぼす格子力の影響 (2007)
  • Author(s): 山本昌孝、早川直紀、村上緑、神山勉
  • Organizer: 日本生物物理学会第45回年会
  • Place of Presentation: 横浜
• [Presentation] スルメイカロドプシンのX線結晶構造解析 (2007)
  • Author(s): 村上緑、神山勉
  • Organizer: 日本生物物理学会第45回年会
  • Place of Presentation: 横浜
• [Presentation] アーキロドプシン-2のM中間体のX線構造解析 (2007)
  • Author(s): 藤井隆道、吉村恵子、村上緑、神山勉
  • Organizer: 日本生物物理学会第45 回年会
  • Place of Presentation: 横浜
• [Presentation] イカロドプシンの結晶構造 (2007)
  • Author(s): 村上緑、神山勉
  • Organizer: 日本結晶学会2007年度年会
  • Place of Presentation: 東京
• [Presentation] バクテリオロドプシンの光反応サイクルに及ぼすXe ガスの影響 (2007)
  • Author(s): 早川直紀、笠原隆史、長谷川大祐、村上緑、神山勉
  • Organizer: 日本結晶学会2007年度年会
  • Place of Presentation: 東京
• [Presentation] バクテリオロドプシンの光誘起構造変化に及ぼす結晶格子力の影響 (2007)
  • Author(s): 山本昌孝、早川直紀、村上緑、神山勉
  • Organizer: 日本結晶学会2007年度年会
  • Place of Presentation: 東京
• [Presentation] Binding of xenon to a hydrophobic cavity in the cytoplasmic half of bacteriorhodopsin & its effect to the absorption kinetics (2007)
  • Author(s): N. Hayakawa, M. Murakami, T. Kasahara, D. Hasegawa and T. Kouyama
  • Organizer: Nagoya International Symposium on Science of Molecular Assembly and Biomolecular Systems
  • Place of Presentation: Naogya
• [Presentation] Decay kinetics of the M intermediate of bR in 3D crystals: Influence of the crystal lattice force (2007)
  • Author(s): M. Yamamoto, N. Hayakawa, M. Murakami and T. Kouyama
  • Organizer: Nagoya International Symposium on Science of Molecular Assembly and Biomolecular Systems, Nagoya
  • Place of Presentation: Naogya
• [Presentation] Crystal Structure of Squid Rhodopsin (2007)
  • Author(s): M. Murakami and T. Kouyama
  • Organizer: International Symposium on RETINAL PROTEINS:EXPERIMENTS AND THEORY
  • Place of Presentation: Bremen, Germany
• [Presentation] Binding of Noble Gases to a Hydrophobic Cavity in the Cytoplasmic Half of Bacteriorhodopsin (2007)
  • Author(s): N. Hayakawa, M. Murakami, T. Kasahara, D. Hasegawa and T. Kouyama
  • Organizer: 2nd International Symposium on Diffraction Structural Biology 2007
  • Place of Presentation: Tokyo
• [Presentation] X-ray crystallographic study of squid rhodopsin (2007)
  • Author(s): M. Murakami and T. Kouyama
  • Organizer: 2nd International Symposium on Diffraction Structural Biology 2007
  • Place of Presentation: Tokyo
• [Presentation] Decay kinetics of the M intermediate of bR in 3D crystals: Influence of the crystal lattice force (2007)
  • Author(s): M. Yamamoto, N. Hayakawa, M. Murakami and T. Kouyama
  • Organizer: 2nd International Symposium on Diffraction Structural Biology 2007
  • Place of Presentation: Tokyo
• [Remarks]
  • URL: http://www.nagoya-u.ac.jp/pdf/research/news/20080515_sci.pdf?20090424
• [Remarks]
  • URL: http://www.spring8.or.jp/ja/current_result/press_release/2008/080515-2
• [Patent(Industrial Property Rights)] イカロドプシン複合体結晶及びその製造方法 (2008)
  • Inventor(s): 神山勉、村上緑
  • Industrial Property Rights Holder: 神山勉、村上緑
  • Industrial Property Number: 2008-058864
  • Filing Date: 2008-03-09
• [Patent(Industrial Property Rights)] 特許権 (2008)
  • Inventor(s): 神山勉、村上緑
  • Industrial Property Rights Holder: 神山勉、村上緑
  • Industrial Property Number: 2008-058864
  • Filing Date: 2008-03-09