Tight binding of inhibitors to bovine bc1 complex is independent of the Rieske protein redox state. Consequences for semiquinone stabilization in the quinol oxidation site.
To determine the effect of the redox state of the Rieske protein on ligand binding to the quinol oxidation site of the bc(1) complex, we measured the binding rate constants (k(1)) for stigmatellin and myxothiazol, at different concentrations of decylbenzoquinone or decylbenzoquinol, in the bovine bc(1) complex with the Rieske protein in the oxidized or reduced state. Stigmatellin and myxothiazol bound tightly and competitively with respect to quinone or quinol, independently of the redox state of the Rieske protein. In the oxidized bc(1) complex, the k(1) values for stigmatellin ( approximately 2.6 x 10(6) m(-1)s(-1)) and myxothiazol ( approximately 8 x 10(5) m(-1)s(-1)), and the dissociation constant (K(d)) for quinone, were similar between pH 6.5 and 9, indicating that ligand binding is independent of the protonation state of histidine 161 of the Rieske protein (pK(a) approximately 7.6). Reduction of the Rieske protein increased the k(1) value for stigmatellin and decreased the K(d) value for quinone by 50%, without modifying the k(1) for myxothiazol. These results indicate that reduction of the Rieske protein and protonation of histidine 161 do not induce a strong stabilization of ligand binding to the quinol oxidation site, as assumed in models that propose the existence of a highly stabilized semiquinone as a reaction intermediate during quinol oxidation.
D000818:Animals; D016227:Benzoquinones / Q000378:metabolism*; D002417:Cattle; D014450:Electron Transport Complex III / Q000037:antagonists & inhibitors* / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000378:metabolism*; D006873:Hydroquinones / Q000378:metabolism*; D007506:Iron-Sulfur Proteins / Q000378:metabolism*; D007700:Kinetics; D008689:Methacrylates; D010084:Oxidation-Reduction; D011090:Polyenes / Q000378:metabolism; D013844:Thiazoles / Q000378:metabolism
D016227:Benzoquinones; D004791:Enzyme Inhibitors; D006873:Hydroquinones; D007506:Iron-Sulfur Proteins; D008689:Methacrylates; D011090:Polyenes; C037989:Rieske iron-sulfur protein; D013844:Thiazoles; C004532:quinone; C030517:myxothiazol; C041573:stigmatellin; D014450:Electron Transport Complex III