Structural analysis of band 3 in the membrane

Research Project

Project/Area Number	23570201
Research Category	Grant-in-Aid for Scientific Research (C)
Allocation Type	Multi-year Fund
Section	一般
Research Field	Biophysics
Research Institution	Okinawa Institute of Science and Technology Graduate University (2013-2014) The Institute of Physical and Chemical Research (2011-2012)
Principal Investigator	HIRAI Teruhisa 沖縄科学技術大学院大学, その他の研究科, 研究員 (10450412)
Co-Investigator(Renkei-kenkyūsha)	HAMASAKI Naotaka 長崎国際大学, 薬学部, 教授 (00091265)
Project Period (FY)	2011-04-28 – 2015-03-31
Project Status	Completed (Fiscal Year 2014)
Budget Amount *help	¥5,200,000 (Direct Cost: ¥4,000,000、Indirect Cost: ¥1,200,000) Fiscal Year 2013: ¥2,080,000 (Direct Cost: ¥1,600,000、Indirect Cost: ¥480,000) Fiscal Year 2012: ¥1,690,000 (Direct Cost: ¥1,300,000、Indirect Cost: ¥390,000) Fiscal Year 2011: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
Keywords	赤血球 / バンド３ / AE1 / 二次元結晶 / 電子線結晶学 / 電子線結晶構造解析 / 膜タンパク質 / 二次元結晶化 / 生体分子 / 蛋白質 / 電子顕微鏡 / 膜輸送体
Outline of Final Research Achievements	We tried to improve the quality of two-dimensional crystal of band 3. We had already got vesicle-like crystals of outward-open band 3 but we tried to get sheet-like crystals to achieve higher resolution. For this purpose, we checked the quality of the sample carefully. We tried to eliminate minor contamination bands that probably corresponded to glycophorin A (GPA) and aquaporin-1. We also checked the stability of purified inward-open band 3 to seek for the other conformation. Besides crystallization efforts of band 3, we tried to purify the band 3 complex. We used Wright monoclonal antibody, which is known to bind to the band 3 and GPA complex, and we were able to get the purified band 3/GPA/Wright monoclonal antibody triple complex.