Characterisation of binding of the methoxyacrylate inhibitors to mitochondrial cytochrome c reductase.
The three E-beta-methoxyacrylate (MOA) inhibitors oudemansin A, strobilurin A and MOA stilbene [3-methoxy-2(2-styrylphenyl)propenic acid-methylester], which differ by more than one order of magnitude in their binding affinity to the mitochondrial ubihydroquinone:cytochrome c oxidoreductase (bc1 complex), bind to a site that is not identical to the binding site for ubihydroquinone, the substrate of the outer ubiquinone reaction site (Qo centre). Although the ubihydroquinone molecule is still bound in the presence of the MOA inhibitors, its electrons cannot be transferred to the iron-sulfur centre. A shift of the relative position of the ubihydroquinone molecule in the reaction centre due to a conformational distortion of cytochrome b induced by the binding of the MOA inhibitor seems to be the reason for the blocked electron transfer. Further analysis shows that ubihydroquinone affects the Kd values of all three MOA inhibitors tested: the values are raised by a constant factor of two, although the inhibitors bind with quite different affinity. The iron-sulfur protein is not involved in the binding of the MOA inhibitors. These results have direct implications for the proper use of MOA inhibitors in experiments designed to analyse the structure/mechanism relationship in cytochrome c reductase. In particular, point mutations recently described in MOA-inhibitor-resistant mutants can no longer be taken to affect necessarily the ubihydroquinone binding site.
Brandt
U
U
Institut für Physikalische Biochemie, Universität München, Federal Republic of Germany.
Schägger
H
H
von Jagow
G
G
D000179:Acrylates / Q000494:pharmacology; D000475:Alkenes / Q000494:pharmacology; D001665:Binding Sites / Q000187:drug effects; D003579:Cytochrome Reductases / Q000037:antagonists & inhibitors*; D003573:Cytochrome b Group / Q000037:antagonists & inhibitors; D004735:Energy Transfer; D005231:Fatty Acids, Unsaturated; D007700:Kinetics; D008689:Methacrylates; D008928:Mitochondria / Q000187:drug effects / Q000201:enzymology*; D009245:NADH Dehydrogenase / Q000037:antagonists & inhibitors*; D010084:Oxidation-Reduction / Q000187:drug effects; D010101:Oxygen Consumption / Q000187:drug effects; D013267:Stilbenes / Q000494:pharmacology*; D000073739:Strobilurins
D000179:Acrylates; D000475:Alkenes; D003573:Cytochrome b Group; D005231:Fatty Acids, Unsaturated; D008689:Methacrylates; D013267:Stilbenes; D000073739:Strobilurins; C056108:3-methoxy-2-(2-styrylphenyl)propenic acid methyl ester; C008556:mucidin; C023324:oudemansin; D003579:Cytochrome Reductases; D009245:NADH Dehydrogenase