The incorporation of reaction centres into membranes from a bacteriochlorophyll-less mutant of Rhodopseudomonas sphaeroides

Abstract

Reaction centres purified from a blue-green mutant R-26 of Rhodopseudomonas sphaeroides can be incorporated into bacteriochlorophyll-less membranes purified from an aerobically-grown bacteriochlorophyll-less mutant 01 of R. sphaeroides. This can be accomplished by raising the temperature of the mixture or by addition of the detergent sodium cholate and its subsequent removal by dilution or dialysis. Optimum conditions for the reconstitution are at 4°C in the presence of 1% cholate and soybean phospholipid (2 : 1, w/w, with membrane protein). Isopycnic sucrose density gradient centrifugation of such preparations shows that reaction centres and light-harvesting pigment-protein complex bind to the membranes. Reconstituted membranes exhibit light-induced steady-state cytochrome absorbance changes resembling those observed in chromatophores prepared from the photosynthetically-grown mutant R-26. The effect on these absorbance changes of varying reaction centre content in the membrane has been studied, and the time course of the interaction between 01 membrane cytochrome c 2 and added reaction centre examined. Cytochrome b photoreduction and cytochrome c 2 photo-oxidation were observed in the reconstituted preparation; each increased following the addition of antimycin A, suggesting that a cyclic light-driven system had been reconstituted.