Mechanism of ubiquinol oxidation by the bc(1) complex: Different domains of the quinol binding pocket and their role in the mechanism and binding of inhibitors

Structures Of mitochondrial ubihydroquinone:cytochrome c oxidoreductase (bc (1) complex) from several animal sources have provided a basis for understa nding the functional mechanism at the molecular level. Using structures of the chicken complex with and without inhibitors, we analyze the effects of mutation on quinol oxidation at the Q(o) site of the complex. We suggest a mechanism for the reaction that incorporates two features revealed by the s tructures, a movement of the iron sulfur protein between two separate react ion domains on cytochrome c(1) and cytochrome b and a bifurcated volume for the Q(o) site. The volume identified by inhibitor binding as the Q(o) site has two domains in which inhibitors of different classes bind differential ly; a domain proximal to heme b(L), where myxothiazole and beta-methoxyacry late(MOA-) type inhibitors bind (class II), and a distal domain close to th e iron sulfur protein docking interface, where stigmatellin and 5-n-undecyl -6-hydroxy-4,7-dioxobenzothiaole (UHDBT) bind (class I). Displacement of on e class of inhibitor by another is accounted for by the overlap of their vo lumes, since the exit tunnel to the lipid phase forces the hydrophobic "tai ls" to occupy common space. We conclude that the site can contain only one "tailed" occupant, either an inhibitor or a quinol or one of their reaction products. The differential sensitivity of strains with mutations in the di fferent domains is explained by the proximity of the affected residues to t he binding domains of the inhibitors. New insights into mechanism are provi ded by analysis of mutations that affect changes in the electron paramagnet ic resonance (EPR) spectrum of the iron sulfur protein, associated with its interactions with the Q(o)-site occupant. The structures show that all int eractions with the iron sulfur protein must occur at the distal position. T hese include interactions between quinone, or class I inhibitors, and the r educed iron sulfur protein and formation of a reaction complex between quin ol and oxidized iron sulfur protein. The step with high activation energy i s after formation of the reaction complex, likely in formation of the semiq uinone and subsequent dissociation of the complex into products. We suggest that further progress of the reaction requires a movement of semiquinone t o the proximal position, thus mapping the bifurcated reaction to the bifurc ated volume. We suggest that such a movement, together with a change in con formation of the site, would remove any semiquinone formed from further int eraction with the oxidized [2Fe-2S] center and also from reaction with O-2 to form superoxide anion. We also identify two separate reaction paths for exit of the two protons released in quinol oxidation.