Purified cytochrome b-c 1 complexes from beef heart mitochondria and Rhodobacter sphaeroides were reconstituted into potassium-loaded asolectin liposomes for studies of the energy-dependent electron transfer reactions within the complexes. Both complexes in a ubiquinone-sufficient state exhibit antimycin-sensitive reduction of cytochromes b (both low and high potential ones) upon induction of a diffusion potential by valinomycin in the presence of ascorbate. Addition of

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... nylenediamine (TMPD) to the ascorbatereduced potassium-loaded asolectin proteoliposomes resulted in reduction of cytochrome b 562 . Upon addition of valinomycin, the induced diffusion potential caused a partial reoxidation of cytochrome b 562 and partial reduction of cytochrome b 566 in beef heart cytochrome b-c 1 complex in the presence of antimycin and/or myxothiazol. Surprisingly, when ubiquinone-depleted beef heart cytochrome b-c 1 complex liposomes were treated under the same conditions, no cytochrome b 566 reduction was observed but only the oxidation of cytochrome b 562 , and the oxidation was not oxygen-dependent. We explain this effect by b 566 , iron-sulfur protein short-circuiting under these conditions, assuming that both antimycin and myxothiazol markedly affect subunit b conformation. The electrochemical midpoint potential of heme b 566 appears to be significantly higher than that of heme b 562 in the presence of myxothiazol, which cannot be accounted for only by the potential-driven electron transfer between these two hemes plus the shift in chemical midpoint potentials caused by myxothiazol. A model for energy coupling consistent with structural findings by Ohnishi et al. (Ohnishi, T., Schagger, H., Meinhardt, S. W., LoBrutto, R., Link, T. A., and von Jagow, G. (1989) J. Biol. Chem. 264, 735-744) is presented. This model is a compromise between pure "redox-loop" and pure "proton-pump" mechanisms. Reoxidation of high potential heme b is observed in an antimycin-or antimycin plus myxothiazol-inhibited, ascorbate plus TMPD-prereduced R. sphaeroides b-c 1 complex, upon membrane potential development, suggesting that a similar electron transfer mechanism is also operating in the bacterial complex.