Equilibrium unfolding of a small low-potential cytochrome, cytochrome c(553) from Desulfovibrio vulgaris

To understand general aspects of stability and folding of c-type cytochrome s, we have studied the folding characteristics of cytochrome C-553 from Des ulfovibrio vulgaris (Hildenborough). This cytochrome is structurally simila r but lacks sequence homology to other heme proteins; moreover, it has an a bnormally low reduction potential. Unfolding of oxidized and reduced cytoch rome C-553 by guanidine hydrochloride (GuHCl) was monitored by circular dic hroism (CD) and Soret absorption; the same unfolding curves were obtained w ith both methods supporting that cytochrome c(553) unfolds by an apparent t wo-state process. Reduced cytochrome C-553 is 7(3) kJ/mol more stable than the oxidized form; accordingly, the reduction potential of unfolded cytochr ome C-553 is 100(20) mV more negative than that of the folded protein. In c ontrast to many other unfolded cytochrome c proteins, upon unfolding at pH 7.0 both oxidized and reduced heme in cytochrome C-553 become high-spin. Th e lack of heme misligation in unfolded cytochrome C-553 implies that its un folded structure is less constrained than those of cytochromes c with low-s pin, misligated hemes.