Conservation of the free energy change of the alkaline isomerization in mitochondrial and bacterial cytochromes c

The thermodynamic parameters of the alkaline transition for oxidized native yeast iso-1 cytochrome c and Rhodopseudomonas palustris cytochrome c2 (cytc2) have been determined through direct electrochemistry experiments carried out at variable pH and temperature and compared to those for horse and beef heart cytochromes c. We have found that both transition enthalpy and entropy are remarkably species dependent, following the order R. palustris cytc2≫ beef (horse) heart cytc > yeast iso-1 cytc. Considering the high homology at the heme–protein interface in the native species, this variability is likely to be mainly determined by differences in the structural and solvation properties and the relative abundance of the various alkaline conformers. Notably, changes in transition enthalpy and entropy among these cytochromes c are compensative and result in small variations in the free energy change of the process (which amounts approximately to +50 kJ mol−1) and consequently in the apparent pKa value. This compensation indicates that solvent reorganization effects play an important role in the thermodynamics of the transition. This mechanism is functional to ensure a relatively high pKa value for the alkaline transition, which is needed to preserve His,Met ligation to the heme iron in cytochrome c at physiological pH and temperature, hence the E° value required for the biological function.