Factors Affecting the Binding of Antimycin A to Complex III of the Mitochondrial Respiratory Chain

Abstract

Abstract The nature of the binding of antimycin A to Complex III of the mitochondrial respiratory chain was investigated according to two procedures. In the first procedure, tritium-labeled antimycin A was extracted from its binding to Complex III either by acetone or by concentrated solutions of taurocholate. Seventy per cent of the complex-bound antimycin A was extracted by either reagent. The acetone-extracted antimycin A was found to be fully inhibitory to the reduced coenzyme Q-cytochrome c reductase activity of Complex III. Moreover, even though most of the antimycin A was extracted from Complex III by taurocholate, its inhibitory effect on the cleavage of the complex was retained. The second procedure utilized enzymically active Complex III as a reagent to detect antimycin A that failed to be bound to, or was released from, an enzymically inactive Complex III. The binding of antimycin A to intact Complex III was found to be essentially irreversible, little or no transfer of antimycin A from antimycin-treated complex to untreated complex being observed. High levels of detergent (3 to 5% taurocholate) in the reaction mixture permitted some transfer of the antimycin A. Conditions that normally caused cleavage of Complex III (i.e. bile salts plus ammonium sulfate, or 1.0 m guanidine), or digestion with trypsin, destroyed the capacity of the complex to bind antimycin A. Reagents that cleaved Complex III despite the treatment of the complex with antimycin A (i.e. 3.0 m guanidine, 0.2 m guanidine plus freezing, or taurocholate plus mersalyl) also caused a release of antimycin A from the complex. Noninhibitory derivatives of antimycin A (i.e. the O-methyl ether of antimycin or deformylantimycin A) offered little or no competition to antimycin A in its binding to Complex III.