Characterization of ATPase Activity of Free and Immobilized Chromatophore Membrane Vesicles of Rhodobacter sphaeroides
2017
Kim, H., Sogang University, Seoul, Republic of Korea | Xiaomeng Tong, Sogang University, Seoul, Republic of Korea | Choi, S., Kyung Hee University, Gyeonggi-do, Republic of Korea | Lee, J.K., Sogang University, Seoul, Republic of Korea
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 (greater than or equal to 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 ㎍ 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 degree Celsius. 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.
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