Structural Studies of Yeast Flavocytochrome b2: Cooperative Roles of the alpha and beta Globules in the Formation of the Flavin-Binding Sites

Abstract

The localization of the heme binding sites on the 2 globular fragments, .alpha. and .beta., of the cleaved form of the flavocytochrome b2 chain was studied. These fragments were partially resolved by means of molecular sieving under denaturing conditions (3 M or 6 M guanidine in the presence of 2-mercaptoethanol). They were then renatured in the presence of excesses of flavin mononucleotide and protoheme. The protoheme was quantitatively bound to the .alpha. subunit, confirming previous findings. The flavin binds neither to .alpha. alone nor to .beta. alone, but only to the reassociated .alpha..beta. protomer. The results are discussed in terms of the possible occurrence of gene fusion in the formation of the complex flavocytochrome chain of this particular L-lactate cytochrome c reductase [EC 1.1.2.3] found specifically in yeasts [bakers].