New evidence for the multiplicity of ubiquinone- and inhibitor-binding sites in the mitochondrial complex I

Determination of the number of ubiquinone- and inhibitor-binding sites in t he mitochonidrial complex I (NADH:ubiquinone oxidoreductase) is a controver sial question with a direct implication for elaborating a suitable model to explain the bioenergetic mechanism of this complicated enzyme. We have use d combinations of both selective inhibitors and common ubiquinone-like subs trates to demonstrate the multiplicity of the reaction centers in the compl ex I in contrast with competition studies that have suggested the existence of a unique binding site for ubiquinone. Our results provide new evidence for the existence of at least two freely exchangeable ubiquinone-binding si tes with different specificity for substrates, as well as for a different k inetic interaction of inhibitors with the enzyme. (C) 2000 Academic Press.