Characterization of ATPase Activity of Free and Immobilized Chromatophore Membrane Vesicles of Rhodobacter sphaeroides

: The intracytoplasmic membrane of Rhodobacter sphaeroides readily vesiculates when cells are lysed. The resulting chromatophore membrane vesicle (CMV) contains the photosynthetic machineries to synthesize ATP by ATPase. The light-dependent ATPase activity of CMV was lowered in the presence of O₂, but the activity increased to the level observed under anaerobic condition when the reaction mixture was supplemented with ascorbic acid (≥0.5 mM). Cell lysis in the presence of biotinyl cap phospholipid (bcp) resulted in the incorporation of bcp into the membrane to form biotinylated CMV (bCMV), which binds to streptavidin resin at a ratio of approximately 24 μg bacteriochlorophyll a/ml resin. The ATPase activity of CMV was not affected by biotinylation, but approximately 30% of the activity was lost by immobilization to resin. Interestingly, the remaining 70% of ATPase activity stayed constant during 7-day storage at 4°C. On the contrary, the ATPase activity of bCMV without immobilization gradually decreased to approximately 40% of the initial level in the same comparison. Thus, the ATPase activity of CMV is sustainable after immobilization, and the immobilized bCMV can be used repeatedly as an ATP generator.