Stability of yeast iso‐1‐ferricytochrome c as a function of pH and temperature

Absorbance‐detected thermal denaturation studies of the C102T variant of Saccharomyces cerevisiae iso‐1‐ferricytochrome c were performed between pH 3 and 5. Thermal denaturation in this pH range is reversible, shows no concentration dependence, and is consistent with a 2‐state model. Values for free energy ( ΔG D ), enthalpy ( ΔH D ), and entropy ( ΔS D ) of denaturation were determined as functions of pH and temperature. The value of ΔG D at 300 K, pH 4.6, is 5.1 ± 0.3 kcal mol −1 . The change in molar heat capacity upon denaturation ( ΔC p ), determined by the temperature dependence of ΔH D as a function of pH (1.37 ± 0.06 kcal mol −1 K −1 ), agrees with the value determined by differential scanning calorimetry. pH‐dependent changes in the Soret region indicate that a group or groups in the heme environment of the denatured protein, probably 1 or both heme propionates, ionize with a p K near 4. The C102T variant exhibits both enthalpy and entropy convergence with a ΔH D of 1.30 kcal mol −1 residue −1 at 373.6 K and a ΔS D of 4.24 cal mol −1 K −1 residue −1 at 385.2 K. These values agree with those for other single‐domain, globular proteins.