Oxidation of cytochromes c and c2 by bacterial photosynthetic reaction centers in phospholipid vesicles. 1. Studies with neutral membranes.
The oxidation of cytochrome c2 by photosynthetic reaction center isolated from Rhodopseudomonas sphaeroides and incorporated into unilamellar phosphatidylcholine vesicles was found to be kinetically similar to that observed earlier for reaction centers in low detergent solution [Overfield, R.E., Wraight, C.A., & DeVault, D. (1979) FEBS Lett. 105, 137-142]. At low ionic strength the kinetics were biphasic. The fast phase indicated the formation of a cytochrome-reaction center complex with an apparent binding constant, KB, of about 10(5) M-1. However, KB decreased dramatically with increasing salt concentration, and no fast oxidation was detectable in 0.1 M NaCl. The slow cytochrome oxidation was first order in both cytochrome and reaction centers and, thus, second order overall. Deviations from theoretical second-order behavior were observed when the rate of the first-order back reaction of the primary photoproducts was significant compared to the cytochrome oxidation. This can cause serious overestimation of the second-order rate constant. The slow oxidation of cytochrome c2 by reaction centers in phosphatidylcholine vesicles exhibited a 40% lower encounter frequency than with the solubilized reaction center. This was attributed to the much lower diffusion coefficient of the reaction center in the vesicle membrane than in solution. No effects of diminished dimensionality were detected with neutral vesicles. An activation energy of 8.0 +/- 0.4 kcal x mol-1 was determined for the slow phase of cytochrome c2 oxidation by reaction centers in solution and in vesicles of several different phosphatidylcholines, including dimyristoylphosphatidylcholine above and below its phase transition temperature. Thus, the physical state of the lipid did not appear to affect any rate-limiting steps leading to cytochrome oxidation. The ionic strength dependence of the slow kinetics of oxidation of cytochromes c and c2 confirmed the electrostatic nature of the cytochrome-reaction center interaction, and the pH dependence indicated the titration of a group or groups, important to this interaction, at pH 9.5.
D003574:Cytochrome c Group / Q000378:metabolism*; D045362:Cytochromes c2; D004579:Electron Transport; D007700:Kinetics; D008081:Liposomes; D009994:Osmolar Concentration; D010084:Oxidation-Reduction; D010713:Phosphatidylcholines; D010788:Photosynthesis; D012242:Rhodobacter sphaeroides / Q000378:metabolism*; D012965:Sodium Chloride / Q000494:pharmacology
D003574:Cytochrome c Group; D008081:Liposomes; D010713:Phosphatidylcholines; D012965:Sodium Chloride; D045362:Cytochromes c2