Effects of mutations on the thermodynamics of a protein folding reaction: Implications for the mechanism of formation of the intermediate and transition states

We have measured changes in heat capacity, entropy, and enthalpy for each s tep in the folding reaction of CD2.d1 and evaluated the effects of core mut ations on these properties. All wild-type and mutant forms fold through a r apidly formed intermediate state that precedes the rate-limiting transition state. Mutations have a pronounced effect on the enthalpy of both the inte rmediate and folded states, but in all cases a compensatory change in entro py results in a small net free-energy change. While the enthalpy change in the folded state can be attributed to a loss of van der Waals interactions, it has already been shown that changes in the stability of the intermediat e are dominated by changes in secondary structure propensity [Lorch et al. (1999) Biochemistry 38, 1377-1385]. It follows that the thermodynamic basis of beta-propensity is enthalpic in origin. The effects of mutations on the enthalpy and entropy of the transition state are smaller than on the groun d states. This relative insensitivity to mutation is discussed in the light of theories concerning the nature of the rate-limiting barrier in folding reactions.