Selective Reduction of Cytochrome C Oxidase with Borohydride.

Since the pioneer studies of Keilin and Hartree (l), considerable evidence has accumulated indicating that cytochrome c oxidase contains two heme moieties that react differently; hence the designation cytochrome U-Q. Reasons for this designat.ion include kinetic studies that show (a) differences in the rate of reduction and oxidation of cytochrome c oxidase measured at the a versus y absorption maxima (2-5), (b) rates of reduction measured at 444 rnp, which indicate the presence of two reducing components (4), and (c) rates of reduction in the presence of CO, which indicate the presence of a fast (444 mM) and a slow (430 rnp) reducing component (6). Recently, Morrison and Horie (7, 8) reported the reduction of the carbonyl group on only the cytochrome a3 heme by pretreatment of the oxidase with borohydride. The cytochrome a heme was not affected, suggesting the selective reducing ability of borohydride toward cytochrome u3. It is the purpose of this paper to show that, contrary to other reports (7,9, lo), borohydride can reduce the iron of the hemes of cytochrome c oxidase. The reduction of iron can be accomplished with or without the concomitant reduction of the carbonyl group of the cytochrome a3 heme. In addition, it is shown that the carbonyl group of cytochrome a was reduced by borohydride only after incubation at alkaline pH.