Structure basis of lipophilic ligand binding of membrane protein involving in lipid mediator biosynthesis

• Project/Area Number: 15H04343
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Structural biochemistry
• Research Institution: Institute of Physical and Chemical Research
• Principal Investigator: Ago Hideo 国立研究開発法人理化学研究所, 放射光科学総合研究センター, 専任研究員 (70360477)
• Project Period (FY): 2015-04-01 – 2018-03-31
• Project Status: Completed (Fiscal Year 2017)
• Budget Amount: ¥16,120,000 (Direct Cost: ¥12,400,000、Indirect Cost: ¥3,720,000); Fiscal Year 2017: ¥3,770,000 (Direct Cost: ¥2,900,000、Indirect Cost: ¥870,000); Fiscal Year 2016: ¥5,720,000 (Direct Cost: ¥4,400,000、Indirect Cost: ¥1,320,000); Fiscal Year 2015: ¥6,630,000 (Direct Cost: ¥5,100,000、Indirect Cost: ¥1,530,000)
• Keywords: Protein crystallography / membrane protein / lipid mediator / 膜タンパク質 / 放射光構造生物 / 脂質メディエータ / X線結晶構造解析 / 放射光 / 蛋白質 / 膜蛋白質

Outline of Final Research Achievements

Structure basis of lipophilic ligand recognition of human membrane proteins involving in lipid mediator biosynthesis was the subject of this study. The membrane proteins utilize hydrophilic GSH and lipophilic arachidonic acid metabolites. In comparison to GSH whose binding architecture was elucidated in detail by X-ray crystallography, limited information about lipophilic ligand was available. This study used two methods to get the lipophilic ligand / protein complex, one was the soaking method on the crystals grown in surfo and the other was the co-crystallization in meso. In addition to making lipophilic ligand / protein complex, a method spreading thinly a highly viscus vehicle in which a large number of micro-crystals were dispersed was developed. Crystal overlap along X-ray beam causes the double hit which makes the diffraction image unavailable. The thin spreading reduces the crystal overlap; thus, resulting in the efficient data collection from micro crystals.

Research Products

• [Journal Article] Protein microcrystallography using synchrotron radiation (2017)
  • Author(s): Yamamoto Masaki、Hirata Kunio、Yamashita Keitaro、Hasegawa Kazuya、Ueno Go、Ago Hideo、Kumasaka Takashi
  • Journal Title: IUCrJ Volume: 4 Issue: 5 Pages: 529-539
  • DOI: 10.1107/s2052252517008193
  • Peer Reviewed / Open Access
• [Journal Article] Development of a dose-limiting data collection strategy for serial synchrotron rotation crystallography (2017)
  • Author(s): Hasegawa Kazuya、Yamashita Keitaro、Murai Tomohiro、Nuemket Nipawan、Hirata Kunio、Ueno Go、Ago Hideo、Nakatsu Toru、Kumasaka Takashi、Yamamoto Masaki
  • Journal Title: Journal of Synchrotron Radiation Volume: 24 Issue: 1 Pages: 29-41
  • DOI: 10.1107/s1600577516016362
  • NAID: 120005947170
  • Peer Reviewed / Open Access
• [Presentation] Fixed target serial crystallography at SACLA (2017)
  • Author(s): Masaki Yamamoto, Hideo Ago, Kunio Hirata, Keitaro Yamashita, Go Ueno, Minoru Kubo, Seiki Baba, Kazuya Hasegawa, Takashi Kumasaka, Atsuhiro Shimada, Kyoko Shinzawa-Itoh, Tomitake Tsukihara, Shinya Yoshikawa, Michihiro Suga, Fusamichi Akita, Jian-Ren Shen
  • Organizer: ACA2017
  • Int'l Joint Research