HPLC purification of a biologically active membrane protein: the reaction center from photosynthetic bacteria.
Reaction centers from Rhodopseudomonas sphaeroides R 26 have been isolated from a crude extract obtained by lauryldimethylamine oxide extraction of chromatophore membranes, by HPLC using a combination of surface-mediated and size exclusion chromatography. The eluted RCs exhibit a normal activity (t 1/2 of the back-reaction is 70 ms) and are recovered in good yield (over 50% based on the activity) and purity (based on the A 280 nm/A 800 nm = 1.30 +/- 0.05 ratio and the characteristic 3 polypeptides SDS-PAGE pattern). The elution time (5-10 mn) is about two orders of magnitude faster than for the classical purification techniques.
D001426:Bacterial Proteins / Q000302:isolation & purification*; D002851:Chromatography, High Pressure Liquid; D003902:Detergents; D004123:Dimethylamines; D006863:Hydrogen-Ion Concentration; D008565:Membrane Proteins / Q000302:isolation & purification*; D009994:Osmolar Concentration; D010777:Photochemistry; D045322:Photosynthetic Reaction Center Complex Proteins; D012242:Rhodobacter sphaeroides / Q000032:analysis*
D001426:Bacterial Proteins; D003902:Detergents; D004123:Dimethylamines; D008565:Membrane Proteins; D045322:Photosynthetic Reaction Center Complex Proteins; C014518:dodecyldimethylamine oxide