ИСТИНА

Succinate:quinone oxidoreductases (SQR) from Bacilli catalyze energetically unfavorable reduction of menaquinone by succinate and this activity is lost upon ΔμН+ dissipation, for example during isolation of leaky membranes from the cells [1]. As shown in [2], tightly coupled membrane vesicles from Bacillus subtilis retain the ability to respire on succinate with the rate approaching that of the whole cells. Here we report that in the tightly coupled vesicles from B. subtilis SQR can be activated by membrane energization by virtue of ATP hydrolysis. Respiratory chain of B. subtilis strain with cytochrome bd as the only terminal oxidase generates ΔμН+ with low efficiency (1H+/e−) and, contrary to the wild type, mutant membranes reveal no succinate oxidase activity even under tightly coupled conditions. Addition of ATP to the inside-out vesicles results, first, in membrane energization and, second, in the ability to oxidize succinate. Subsequent uncoupling inhibits succinate respiration completely. On the other hand, neither ATP-driven energization nor uncoupling affects the reversed, fumarate reductase, activity of the membranes. The latter fact is in accordance with our finding that the menaquinol: fumarate reductase activity of B. subtilis membranes is not coupled to ΔμН+ generation. In order to identify the potential ΔμН+-sensitive step(s) of the succinate:menaquinone reaction, the redox state of the di-heme cytochrome b forming the membrane anchor part of SQR was assayed. To this end, membranes were equilibrated with the succinate/fumarate redox couple so that the reduction of the high (bH) and low (bL) potential hemes were near 100% and 50%, respectively. The membranes were then energized by addition of ATP. The energization did not result in any change in the reduction level of the hemes, as well as subsequent uncoupling. Presumably, the effect of the ATPase-imposed Δψ on electron distribution among the redox centers of SQR is electrically compensated by intraprotein movement of positive charges (probably, protons), which could be a constituent part of the catalytic