STRUCTURE-BASED STATISTICAL THERMODYNAMIC ANALYSIS OF T4 LYSOZYME MUTANTS - STRUCTURAL MAPPING OF COOPERATIVE INTERACTIONS

The recent development of a structural parameterization of the energet ics of protein folding has permitted the incorporation of the function s that describe the enthalpy, entropy and heat capacity changes, i.e. the individual components of the Gibbs energy, into a statistical ther modynamic formalism that describes the distribution of conformational states under equilibrium conditions. The goal of this approach is to c onstruct with the computer a large ensemble of conformational states, and then to derive the most probable population distribution, i.e. the distribution of states that best accounts for a wide array of experim ental observables. This analysis has been applied to four different mu tants of T4 lysozyme (S44A, S44G, V131A, V131G). It is shown that the structural parameterization predicts well the stability of the protein and the effects of the mutations. The entire set of folding constants per residue has been calculated for the four mutants. In all cases, t he effect of the mutations propagates beyond the mutation site itself through sequence and three-dimensional space. This phenomenon occurs d espite the fact that the mutations are at solvent-exposed locations an d do not directly affect other interactions in the protein. These resu lts suggest that single amino acid mutations at solvent-exposed locati ons, or other locations that cause a minimal perturbation, can be used to identify the extent of cooperative interactions. The magnitude and extent of these effects and the accuracy of the algorithm can be test ed by means of NMR-detected hydrogen exchange.