Challenges for the structure determination of GPCR active conformation

• Project/Area Number: 23657074
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• Research Field: Structural biochemistry
• Research Institution: Kyoto University
• Principal Investigator: SHIMAMURA Tatsuro 京都大学, 大学院・医学研究科, 特定講師 (90391979)
• Co-Investigator(Renkei-kenkyūsha): OGASAWARA Satoshi 京都大学, 医学研究科, 研究員 (30546685) ; ASADA Hidetsugu 京都大学, 医学研究科, 研究員 (20399041)
• Project Period (FY): 2011 – 2012
• Project Status: Completed (Fiscal Year 2012)
• Budget Amount: ¥3,900,000 (Direct Cost: ¥3,000,000、Indirect Cost: ¥900,000); Fiscal Year 2012: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2011: ¥2,470,000 (Direct Cost: ¥1,900,000、Indirect Cost: ¥570,000)
• Keywords: X線結晶解析 / 受容体 / GPCR / 膜タンパク質 / 構造解析 / G蛋白質共役型受容体 / X線結晶構造解析

Research Abstract

In order to understand the GPCR-mediated signal transduction mechanisms, it is essential to determine the GPCR structures in the active and inactive conformations. We published the structure of Histamine H1 receptor in complex with an inverse agonist doxepin. The structure revealed the reason why doxepin shows high affinities to aminergic receptors. We also published the structure of Adenosine A2a receptor in the inactive conformation. For crystallization, the receptor was stabilized in the inactive conformation by the binding of Fab fragment. The antibody that stabilizes the receptor in the active conformation was successfully obtained. This antibody will be useful for the determination of the Adenosine A2a receptor structure in the active conformation.

Research Products

• [Journal Article] Structural biology and molecular targets (2012)
  • Author(s): Shimamura T, Iwata S.
  • Journal Title: Nihon Rinsho Volume: Nov;70 Suppl 8 Pages: 316-20
• [Journal Article] Structure of histamine H1 receptor (2012)
  • Author(s): Shimamura T
  • Journal Title: Seikagaku. Volume: 84(9) Pages: 772-6
  • NAID: 10031122382
• [Journal Article] Conformational transition of the lid helix covering the proteaseactive site is essential for the ATP-dependent protease activity of FtsH. (2012)
  • Author(s): Suno R, Shimoyama M, Abe A, Shimamura T, Shimodate N, Watanabe YH, Akiyama Y, Yoshida M.
  • Journal Title: FEBS Lett Volume: 586(19) Pages: 3117-21
  • Peer Reviewed
• [Journal Article] Platform for the rapid construction and evaluation of GPCRs for crystallography in Saccharoroyces cerevisiae. (2012)
  • Author(s): Shiroishi M., et al.
  • Journal Title: Micorb, Cell Fact. Volume: 11 Issue: 1 Pages: 78-78
  • DOI: 10.1186/1475-2859-11-78
  • Peer Reviewed
• [Journal Article] G protein-coupled receptor inactivation by an allosteric inverse-agonist antibody (2012)
  • Author(s): Hino T., et al
  • Journal Title: Nature Volume: 482 Issue: 7384 Pages: 237-40
  • DOI: 10.1038/nature10750
  • Peer Reviewed
• [Journal Article] Conformational transition of the lid helix covering the protease active site is essential for the ATP-dependent protease activity of FtsH. (2012)
  • Author(s): Suno R. et al.
  • Journal Title: FEBS Lett Volume: 586 Issue: 19 Pages: 3117-21
  • DOI: 10.1016/j.febslet.2012.07.069
  • Peer Reviewed
• [Journal Article] Structure of histamine H1 receptor. (2012)
  • Author(s): Shimamura T.
  • Journal Title: Seikagaku. Volume: 84 Pages: 772-6
  • NAID: 10031122382
• [Journal Article] Structural biology and molecular targets. (2012)
  • Author(s): Shimamura T.
  • Journal Title: Nihon Rinsho. Volume: 70 Pages: 316-20
• [Journal Article] The Na+-hydantoin membrane transport protein, Mhp1. (2012)
  • Author(s): Jackson SM.et al.
  • Journal Title: Encyclopedia of Biophysics. Volume: - Pages: 1514-1521
• [Journal Article] 創薬ターゲット膜蛋白質の立体構造解析 (2012)
  • Author(s): 島村達郎
  • Journal Title: 内分泌・糖尿病・代謝内科 Volume: 35 Pages: 557-564
• [Journal Article] Crystal structure-based virtual screening for fragment-like ligands of the human histamine H(1) receptor. (2011)
  • Author(s): de Graaf C, Kooistra AJ, Vischer HF, Katritch V, Kuijer M, Shiroishi M, Iwata S, Shimamura T, Stevens RC, de Esch IJ, Leurs R.
  • Journal Title: J Med Chem. Volume: 54(23) Issue: 23 Pages: 8195-206
  • DOI: 10.1021/jm2011589
  • Peer Reviewed
• [Journal Article] Production of the stable human histamine H(1) receptor in Pichia pastoris for structural determination (2011)
  • Author(s): Shiroishi M, Kobayashi T, Shimamura T., et al
  • Journal Title: Methods Volume: 55 Issue: 4 Pages: 281-286
  • DOI: 10.1016/j.ymeth.2011.08.015
  • Peer Reviewed
• [Journal Article] Structure ofthe human histamine H1 receptor complexwith doxepin. (2011)
  • Author(s): Shimamura, T., Shiroishi, M, Weyand, S., Tsujimoto, H., Winter, G., Katritch, V., Abagyan, R., Cherezov, V., Liu, W., Han, G. W., Kobayashi, T., Stevens, R. C. and Iwata, S
  • Journal Title: Nature Volume: 475 Issue: 7354 Pages: 65-70
  • DOI: 10.1038/nature10236
  • Peer Reviewed
• [Journal Article] Cloning, expression and purification of the anion exchanger 1 homologue from the basidiomycete Phanerochaete chrysosporium (2011)
  • Author(s): Tokuda N, Igarashi K, Shimamura T(co-1st author), Yurugi-Kobayashi T, Shiroishi M, Ito K, Sugawara T, Asada H, Murata T, Nomura N, Iwata S, Kobayashi T.
  • Journal Title: Protein. Expr. Purif. Volume: 79(1) Issue: 1 Pages: 81-7
  • DOI: 10.1016/j.pep.2011.04.006
  • NAID: 120003255719
  • Peer Reviewed
• [Journal Article] ヒスタミンH1受容体の立体構造解明と創薬への期待 (2011)
  • Author(s): 島村達郎、岩田想
  • Journal Title: 日本医事新報 Volume: 4564 Pages: 50-51
• [Presentation] ヒスタミンH1受容体の構造により解明された抗ヒスタミン薬選択性機構 (2012)
  • Author(s): 島村 達郎
  • Organizer: 第34回生体膜と薬物の相互作用シンポジウム
  • Place of Presentation: 京都
  • Year and Date: 2012-11-15
• [Presentation] キュービックフェーズ法による膜タンパク質の結晶化 (2012)
  • Author(s): 島村 達郎
  • Organizer: 蛋白研セミナー
  • Place of Presentation: 大阪
  • Year and Date: 2012-09-21
• [Presentation] 膜蛋白質の結晶化 (2012)
  • Author(s): 島村 達郎
  • Organizer: 第12回 日本蛋白質科学会
  • Place of Presentation: 名古屋
  • Year and Date: 2012-06-20
• [Presentation] ヒト由来ヒスタミンH1受容体の立体構造 (2012)
  • Author(s): 島村 達郎
  • Organizer: 第9回 GPCR研究会
  • Place of Presentation: 東京
  • Year and Date: 2012-05-11
• [Presentation] Structural studies of human G-protein coupled receptors (2012)
  • Author(s): Shimamura T.
  • Organizer: 日本農芸化学会2012 年度大会
  • Place of Presentation: 京都
  • Year and Date: 2012-03-25
• [Presentation] 微結晶を用いた創薬ターゲット膜タンパク質の構造解析とSACLAの可能性 (2012)
  • Author(s): 島村 達郎
  • Organizer: 第25回日本放射光学会年会放射光科学合同シンポジウム
  • Place of Presentation: 鳥栖
  • Year and Date: 2012-01-09
• [Presentation] 膜蛋白質の結晶化 (2012)
  • Author(s): 島村達郎
  • Organizer: 第12回 日本蛋白質科学会(招待講演)
  • Place of Presentation: 名古屋
• [Presentation] Structural studies of human G-protein coupled receptors. (2012)
  • Author(s): Shimamura T.
  • Organizer: 日本農芸化学会2012年度大会(招待講演)
  • Place of Presentation: 京都
• [Presentation] 微結晶を用いた創薬ターゲット膜タンパク質の構造解析とSACLAの可能性 (2012)
  • Author(s): 島村達郎
  • Organizer: 第25回日本放射光学会年会放射光科学合同シンポジウム(招待講演)
  • Place of Presentation: 鳥栖
• [Presentation] ヒスタミンH1 受容体の立体構造から分かった抗ヒスタミン薬選択性の分子機構 (2011)
  • Author(s): 島村 達郎
  • Organizer: 第18回創薬情報研究会
  • Place of Presentation: 大阪
  • Year and Date: 2011-12-22
• [Presentation] Structural basis of secondary active transport in the sodium-hydantoin transporter Mhp1. (2011)
  • Author(s): Shimamura T.
  • Organizer: The JSPS/iCeMS International Symposium.
  • Place of Presentation: Kyoto, Japan
  • Year and Date: 2011-11-17
• [Presentation] ヒト由来ヒスタミン受容体H1 の構造解析戦略 (2011)
  • Author(s): 島村 達郎
  • Organizer: 第84回日本生化学会大会
  • Place of Presentation: 京都
  • Year and Date: 2011-09-24
• [Presentation] ヒダントイン輸送体Mhp1の基質輸送中の動き (2011)
  • Author(s): 島村 達郎, 岩田想
  • Organizer: 生理研研究会「作動中の膜機能分子の姿を捉える - 静止画から動画へ -」
  • Place of Presentation: 岡崎
  • Year and Date: 2011-09-08
• [Presentation] Structure of the human histamine H1 receptor with doxepin. (2011)
  • Author(s): Shimamura T.
  • Organizer: International Union of Crystallography, XXII Congress and General Assembly.
  • Place of Presentation: Madrid, Spain.
  • Year and Date: 2011-08-29
• [Presentation] Structure of the human histamine H1 receptor with doxepin. (2011)
  • Author(s): Shimamura T. et al.
  • Organizer: International Union of Crystallography, XXII Congress and General Assembly.(招待講演)
  • Place of Presentation: マドリッド（スペイン）
• [Presentation] ヒト由来ヒスタミン受容体H1の構造解析戦略 (2011)
  • Author(s): 島村達郎ら
  • Organizer: 第84回日本生化学会大会(招待講演)
  • Place of Presentation: 京都
• [Presentation] ヒスタミンH１受容体の構造により解明された抗ヒスタミン薬選択性機構
  • Author(s): 島村 達郎
  • Organizer: 第34回生体膜と薬物の相互作用シンポジウム
  • Place of Presentation: 京都
  • Invited
• [Presentation] 膜蛋白質の結晶化
  • Author(s): 島村 達郎
  • Organizer: 第12回 日本蛋白質科学会
  • Place of Presentation: 名古屋
  • Invited
• [Presentation] ヒト由来ヒスタミンH１受容体の立体構造
  • Author(s): 島村 達郎
  • Organizer: 第９回 GPCR研究会
  • Place of Presentation: 東京
  • Invited
• [Remarks]
  • URL: http://www.kyoto-u.ac.jp/ja/news_data/h/h1/news6/2010/100423_1.htm
• [Remarks]
  • URL: http://www.jst.go.jp/pr/announce/20110623/index.html
• [Remarks]
  • URL: http://www.kyoto-u.ac.jp/ja/news_data/h/h1/news6/2011/110623_1.htm
• [Patent(Industrial Property Rights)] アゴニストの親和性を亢進する抗ヒトアデノシンA2a受容体モノクローナル抗体 (2012)
  • Inventor(s): 小笠原諭 日野智也 島村達郎荒川孝俊 万木貴美 寿野千代 村田武士小林拓也 岩田想
  • Industrial Property Number: 2012-049812
  • Filing Date: 2012-03-06
• [Patent(Industrial Property Rights)] アゴニストの親和性を亢進する抗ヒトアデノシンA2a受容体モノクローナル抗体 (2012)
  • Inventor(s): 小笠原諭, 島村達郎 ら
  • Industrial Property Rights Holder: （独）科学技術振興機構
  • Industrial Property Number: 2012-049812
  • Filing Date: 2012