Interaction of drugs with a model membrane protein. Effects of local anesthetics on electron transfer and hydrogen ion uptake in ionophore stimulated cytochrome oxidase proteoliposomes

Abstract

Cytochrome oxidase extracted from beef heart was incorporated into vesicles composed of soy bean phospholipids (asolectin). The oxidation of externally added cytochrome c by such vesicles is associated with proton uptake from the external medium. The rates of both cytochrome c oxidation and proton uptake were stimulated by addition of ionophores such as trifluoromethoxy carbonyl cyanide phenylhydrazone (FCCP), nigericin and valinomycin. These agents probably dissipate pH and/or electrical potential gradients which develop as a result of enzyme activity and which have a "restraining" influence on the turnover of the oxidase. Local anesthetics inhibited oxidase activity but had a much greater effect on the stimulated (ionophore-treated) than the unstimulated enzyme. In addition, pretreating proteoliposomes with local anesthetics completely prevented the stimulating effects of these ionophores. Based on this and previous studies, a model was developed in which local anesthetics interacted with the phospholipid component of the oxidase complex resulting in reduced internal electron transfer and dissociation of the oxidase from the regulatory role of the proton gradient.