Binding of MOA-stilbene to the mitochondrial cytochrome bc1 complex is affected by the protonation state of a redox-Bohr group of the 'Rieske' iron-sulfur protein.

MOA-stilbene is a specific inhibitor of the ubihydroquinone oxidation center (center P or o) of cytochrome bc1 complex. Binding of this inhibitor does not require the 'Rieske' iron-sulfur protein, but is affected by the redox-state of the cytochrome bc1 complex. We have analyzed the pH dependence of the apparent dissociation constant for MOA-stilbene. A 2.5 fold change in affinity between pH 6.0 and 9.5 was observed for oxidized bovine cytochrome bc1 complex. The pH profile could be simulated by assuming a single protonable group with pKA = 7.7. This pKA was not observed after partial or complete reduction of the enzyme or after removal of the iron-sulfur protein. We conclude that this protonable group was identical to the redox-Bohr group with the same pKA that has been reported to be associated with the 'Rieske' iron-sulfur cluster. Fully reduced cytochrome bc1 complex exhibited an additional binding site for MOA-stilbene. As this second binding site was abolished by the center P inhibitor stigmatellin, but not by antimycin, an inhibitor of ubiquinone reduction at center N, we conclude that it is also located at center P.