Equilibrium and kinetic studies of unfolding of homologous cytochromes c.
Extensive investigations of the unfolding equilibria and kinetics of oxidized and reduced cytochromes c are reported. It is found that all cytochromes c have similar unfolding free energies (deltaGD = 7 +/- 1 kcal/mol). Differences among species do not correlate in any way with the metabolic differences among species. The stabilization of cytochrome c on reduction is estimated at 1.1 kcal/mol. Stability differences among species are mirrored in their denaturation kinetics. For cytochrome c (III), the unfolding exhibits multiple phases. The rate constants for the two observable phases both change by a factor of 3 between horse cytochrome c (III) and cow cytochrome c (III). On reduction, all unfolding appears to occur in a single step. The rate of this unfolding still varies between species, however, the results can be accommodated to a sequential model, with some assumptions. The observations are consistent with chain reversal occurring at an early stage in the reaction and suggest that previously observed rapid processes may be ligand exchange processes.