5-n-undecyl-6-hydroxy-4-7-dioxobenzothiazole and 3-methoxy-2-(2-styrylphenyl)propenic-acid-methyl-ester

Structural analysis of the bc(1) complex suggests that the extra membrane domain of iron-sulfur protein (ISP) undergoes substantial movement during the catalytic cycle. Binding of Qo site inhibitors to this complex affects the mobility of ISP. Taking advantage of the difference in the pH dependence of the redox midpoint potentials of cytochrome c(1) and ISP, we have measured electron transfer between the [2Fe-2S] cluster and heme c(1) in native and inhibitor-treated partially reduced cytochrome bc(1) complexes. The rate of the pH-induced cytochrome c(1) reduction can be estimated by conventional stopped-flow techniques (t1/2, 1-2 ms), whereas the rate of cytochrome c(1) oxidation is too high for stopped-flow measurement. These results suggest that oxidized ISP has a higher mobility than reduced ISP and that the movement of reduced ISP may require an energy input from another component. In the 5-n-undecyl-6-hydroxy-4,7-dioxobenzothiazole (UHDBT)-inhibited complex, the rate of cytochrome c(1) reduction is greatly decreased to a t1/2 of approximately 2.8 s. An even lower rate is observed with the stigmatellin-treated complex. These results support the idea that UHDBT and stigmatellin arrest the [2Fe-2S] cluster at a fixed position, 31 A from heme c(1), making electron transfer very slow.

Topics: Animals; Cattle; Cytochromes c1; Electron Transport; Electron Transport Complex III; Heme; Hydrogen-Ion Concentration; Iron; Iron-Sulfur Proteins; Oxidation-Reduction; Polyenes; Stilbenes; Sulfur; Thiazoles; Time Factors

Native structures of ubihydroquinone:cytochrome c oxidoreductase (bc(1) complex) from different sources, and structures with inhibitors in place, show a 16-22 A displacement of the [2Fe-2S] cluster and the position of the C-terminal extrinsic domain of the iron sulfur protein. None of the structures shows a static configuration that would allow catalysis of all partial reactions of quinol oxidation. We have suggested that the different conformations reflect a movement of the subunit necessary for catalysis. The displacement from an interface with cytochrome c(1) in native crystals to an interface with cytochrome b is induced by stigmatellin or 5-n-undecyl-6-hydroxy-4,7-dioxobenzothiazole (UHDBT) and involves ligand formation between His-161 of the [2Fe-2S] binding cluster and the inhibitor. The movement is a rotational displacement, so that the same conserved docking surface on the iron sulfur protein interacts with cytochrome c(1) and with cytochrome b. The mobile extrinsic domain retains essentially the same tertiary structure, and the anchoring N-terminal tail remains in the same position. The movement occurs through an extension of a helical segment in the short linking span. We report details of the protein structure for the two main configurations in the chicken heart mitochondrial complex and discuss insights into mechanism provided by the structures and by mutant strains in which the docking at the cytochrome b interface is impaired. The movement of the iron sulfur protein represents a novel mechanism of electron transfer, in which a tethered mobile head allows electron transfer through a distance without the entropic loss from free diffusion.

Topics: Amino Acid Sequence; Animals; Anti-Bacterial Agents; Binding Sites; Chickens; Computer Simulation; Crystallography; Cytochrome b Group; Electron Transport Complex III; Enzyme Inhibitors; Iron-Sulfur Proteins; Ligands; Mitochondria, Heart; Molecular Sequence Data; Mutation; Oxidation-Reduction; Polyenes; Protein Engineering; Protein Structure, Secondary; Sequence Alignment; Stilbenes; Thiazoles; Ubiquinone