STABILITY EFFECTS OF INCREASING THE HYDROPHOBICITY OF SOLVENT-EXPOSEDSIDE-CHAINS IN STAPHYLOCOCCAL NUCLEASE

A total Of fifty single site surface phenylalanine substitution mutant s have been made in the model protein staphylococcal nuclease. The fif ty residues that were replaced with phenylalanine were chosen to give a broad sampling of solvent accessibility, secondary structure, and ba ckbone conformations. The change in the stability of each mutant prote in relative to wild type was measured by guanidine hydrochloride denat uration. These results were compared to previous results obtained when these same sites were substituted with an alanine and a glycine. By t his means, changes in the stability due to the loss of interactions of the wild-type side chain can be separated from the effects of introdu cing the bulky, hydrophobic phenylalanine in these solvent-exposed pos itions. In general, our results agree with the conventional wisdom tha t placing a hydrophobic residue in a solvent-exposed position is desta bilizing in most cases, but less destabilizing than most changes in th e hydrophobic core of the protein. However, the degree to which a hydr ophobic surface substitution destabilizes or stabilizes a globular pro tein is highly context-dependent, with some mutations being as destabi lizing as those in the core. This indicates that steric and packing co nsiderations are also important on the surface of a globular protein b ut generally are not as important as in the interior. No evidence for the widespread occurrence of the so-called reverse hydrophobic effect at solvent-exposed sites was found. In addition, this survey of numero us sites suggests that previous measurements of alpha-helix ''propensi ties'' often seriously underestimate the importance of the environment of the side chain.