ISOFORMS OF YEAST CYTOCHROME-C-OXIDASE SUBUNIT-V AFFECT THE BINUCLEARREACTION-CENTER AND AFTER THE KINETICS OF INTERACTION WITH THE ISOFORMS OF YEAST CYTOCHROME-C

Subunit V, one of the nuclear coded subunits of yeast cytochrome c oxi dase, has two isoforms, Va and Vb. These alter the in vivo intramolecu lar rates of electron transfer within the holoenzyme (Waterland, R. A. , Basu, A., Chance, B., and Poyton, R. O. (1991) J. Biol. Chen. 266, 4 180-4186). The isozyme with Vb has a higher turnover rate and a higher intramolecular transfer rate than the isozyme with Va. To determine h ow these isoforms affect catalysis, we have examined their effects on the binuclear reaction center and on the interaction between cytochrom e c oxidase and the two isoforms, iso-1 and iso-2, of yeast cytochrome c. Infrared spectroscopy of carbon monoxide liganded to heme a(3) has revealed a single conformer for the binuclear reaction center in the isozyme with Vb but two discrete conformers in the isozyme with Va. Th e kinetics of interaction for all four pairwise combinations of isozym es with each subunit V isoform and the two cytochrome c isoforms are b iphasic, with high and low affinity electron transfer reactions. In ge neral, the isoforms of cytochrome c and subunit V do not alter the K-m but do affect the TNmax. The TNmax for isozymes carrying Vb are highe r at both high and low affinity sites for each cytochrome c isoform. I so-1-cytochrome c supports a higher TNmax than Iso-2-cytochrome c. Sur prisingly, the combinatorial effect of both sets of isoforms on TNmax is minimized with the pairs of isoforms (iso-1-cytochrome c and subuni t Va or iso-2 and subunit Vb) that are co-expressed in cells. Together , these findings support the conclusion that the subunit V isoforms mo dulate catalysis and suggest that they do so by affecting the environm ent or structure of the binuclear reaction center. They also suggest t hat the coexpression of the two cytochrome c isoforms with two subunit V isoforms serves to minimize differences in electron transfer rates brought about by the subunit V isoforms.