| Title: | A class-A GPCR solubilized under high hydrostatic pressure retains its ligand binding ability |
| Author(s): | Katayama Y; Suzuki T; Ebisawa T; Ohtsuka J; Wang S; Natsume R; Lo YH; Senda T; Nagamine T; Hull JJ; Matsumoto S; Nagasawa H; Nagata K; Tanokura M; |
| Address: | "Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan. Department of Green and Sustainable Chemistry, School of Engineering, Tokyo Denki University, 5 Senju Asahi-cho, Adachi-ku, Tokyo 120-8551, Japan. Structural Biology Research Center, Photon Factory, Institute of Materials Structure Science, High Energy Accelerator Research Organization (KEK), 1-1 Oho, Tsukuba, Ibaraki 305-0801, Japan. Molecular Entomology Laboratory, RIKEN Advanced Science Institute, Wako, Saitama 351-0198, Japan. USDA-ARS Arid Land Agricultural Research Center, Maricopa, AZ, USA. Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan; College of Life Sciences, Zhejiang University, Hangzhou, Zhejiang 310058, PR China. Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan. Electronic address: aknagata@mail.ecc.u-tokyo.ac.jp. Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan. Electronic address: amtanok@mail.ecc.u-tokyo.ac.jp" |
| DOI: | 10.1016/j.bbamem.2016.06.012 |
| ISSN/ISBN: | 0006-3002 (Print) 0006-3002 (Linking) |
| Abstract: | "The effect of high hydrostatic pressure (HHP) on the solubilization of a class-A G protein-coupled receptor, the silkmoth pheromone biosynthesis-activating neuropeptide receptor (PBANR), was investigated. PBANR was expressed in expresSF+ insect cells as a C-terminal fusion protein with EGFP. The membrane fraction was subjected to HHP treatment (200MPa) at room temperature for 1-16h in the presence of 0-2.0% (w/v) n-dodecyl-beta-D-maltopyranoside (DDM). The solubilization yield of PBANR-EGFP in the presence of 0.6% (w/v) DDM increased to ~1.5-fold after 1h HHP treatment. Fluorescence-detection size-exclusion chromatography demonstrated that the PBANR-EGFP ligand binding ability was retained after HHP-mediated solubilization. The PBANR-EGFP solubilized with 1.0% DDM under HHP at room temperature for 6h retained ligand binding ability, whereas solubilization in the absence of HHP treatment resulted in denaturation" |
| Keywords: | "Animals Bombyx/*chemistry/genetics Green Fluorescent Proteins/chemistry/genetics Hydrostatic Pressure Insect Proteins/*chemistry/genetics Protein Stability Receptors, Pheromone/*chemistry/genetics Recombinant Fusion Proteins/chemistry/genetics G protein-c;" |
| Notes: | "MedlineKatayama, Yukie Suzuki, Tatsuya Ebisawa, Tatsuki Ohtsuka, Jun Wang, Shipeng Natsume, Ryo Lo, Yu-Hua Senda, Toshiya Nagamine, Toshihiro Hull, J Joe Matsumoto, Shogo Nagasawa, Hiromichi Nagata, Koji Tanokura, Masaru eng Research Support, Non-U.S. Gov't Netherlands 2016/06/28 Biochim Biophys Acta. 2016 Sep; 1858(9):2145-2151. doi: 10.1016/j.bbamem.2016.06.012. Epub 2016 Jun 21" |
