A thermodynamic study of mutant forms of Streptomyces subtilisin inhibitor. III. Replacements of a hyper-exposed residue, Met73.

Amino acid replacements of Met73, which is hyper-exposed in the native structure, with Asp, Glu, Lys, Gly, Ala, Val, Leu or Ile changed the stability of Streptomyces subtilism inhibitor by +1.60 to -0.94 kcal/mol-1 in free energy at 82.21 degrees C at pH 7.0, with higher hydrophobicity of the replacing amino acid side-chain showing a correlation with lower stability. Thermodynamic parameters obtained from detailed calorimetric analyses gave a nearly proportional relation between the entropy change and the enthalpy change of denaturation, i.e. T delta delta S degrees = -0.03 (+/- 0.11) + 1.14 (+/- 0.03) delta delta H, with the linear correlation coefficient 0.996 for 18 data points. This proportionality observed uniquely for the substitutions at position 73 was caused primarily by the water/side-chain interaction, or hydration effect, which can account for the majority of the changes in enthalpy and entropy induced by the mutations.