CAN ONE PREDICT PROTEIN STABILITY - AN ATTEMPT TO DO SO FOR RESIDUE-133 OF T4 LYSOZYME USING A COMBINATION OF FREE-ENERGY DERIVATIVES, PROFEC, AND FREE-ENERGY PERTURBATION-METHODS

Free energy derivatives, pictorial representation of free energy chang es (PROFEC) and free energy perturbation methods were employed to sugg est the modifications that may improve the stability of a mutant T4 ly sozyme with a S-2-amino-3-cyclopentylpropanoic acid residue (Cpe) at p osition 133. The free energy derivatives and PROFEC methods were used to locate promising sites where modifications may be introduced. The e ffects of several candidate modifications on the enzyme's stability we re analyzed by the free energy perturbation method. We found that this scheme is able to effectively suggest modifications that may increase the enzyme's stability. The modifications investigated are the introd uction of a methyl, a tert-butyl or a trifluoromethyl group at the C-e psilon 2 position and a cyclopropyl group between the C-delta 2 and C- epsilon 2 position on the cyclopentyl ring. The stereochemistry of the introduced groups (in the alpha or beta configurations) was studied. Our calculations predict that the introduction of a methyl group in th e a! configuration or a cyclopropyl group in the beta configuration wi ll increase the stability of the enzyme; the introduction of the two g roups in the other configurations and the other modifications will dec rease the stability of the enzyme. The results indicate that packing i nteractions can strongly influence the stability of the enzyme. (C) 19 98 Wiley-Liss, Inc.