Reaction center and UQH2:cyt c2 oxidoreductase act as independent enzymes in Rps. sphaeroides.

Turnover of the ubiquinol oxidizing site of the UQH2:cyt c2 oxidoreductase (b/c1 complex) of Rps. sphaeroides can be assayed by measuring the rate of reduction of cyt b561 in the presence of antimycin (AA). Oxidation of ubiquinol is a second-order process, with a value of k2 of about 3 X 10(5)M-1. The reaction shows saturation at high quinol concentrations, with an apparent Km of about 6-8mM (with respect to the concentration of quinol in the membrane). When the quinone pool is oxidized before illumination, reduction of the complex shows a substantial lag (about 1 ms) after a flash, indicating that the quinol produced as a result of the photochemical reactions is not immediately available to the complex. We have suggested that the lag may be due to several factors, including the leaving time of the quinol from the reaction center, the diffusion time to the complex, and the time for the head group to cross the membrane. We have suggested a minimal value for the diffusion coefficient of ubiquinone in the membrane (assuming that the lag is due entirely to diffusion) of about 10(-9) cm-2 sec-1. The lag is reduced to about 100 microseconds when the pool is significantly reduced, showing that quinol from the pool is more rapidly available to the complex than that from the reaction center. With the pool oxidized, similar kinetics are seen when the reduction of cyt b561 occurs through the AA-sensitive site (with reactions at the quinol oxidizing site blocked by myxothiazol).(ABSTRACT TRUNCATED AT 250 WORDS)