Are buried salt bridges important for protein stability and conformational specificity?

Abstract

The side chains of Arg 31, Glu 36 and Arg 40 in Arc repressor form a buried salt-bridge triad. The entire salt-bridge network can be replaced by hydrophobic residues in combinatorial randomization experiments resulting in active mutants that are significantly more stable than wild type. The crystal structure of one mutant reveals that the mutant side chains pack against each other in an otherwise wild-type fold. Thus, simple hydrophobic interactions provide more stabilizing energy than the buried salt bridge and confer comparable conformational specificity.

Effects of salt bridges on protein structure and design

“…Moreover, charges involved in salt bridges in the ground state were generally, but not always, more effective in reducing degeneracy than those that were not. Together with accumulating evidence that buried polar and charged interactions tend to destabilize folded proteins relative to hydrophobic groups of similar size and shape (Hendsch & Tidor, 1994;Yang & Honig, 1995;Waldburger et al, 1995;Wang et al, 1996;Wimley et al, 1996), this suggests there is, in general, a trade-off between stability and specificity [i.e., the uniqueness of the native fold (Lattman & Rose, 1993)]. The discovery of particular constellations of interactions that impart stability and specificity simultaneously would thus be extremely valuable for protein molecular engineering.…”

“…Studies of "compound" and "cooperative" interactions have expanded our understanding beyond simply excluded volume, dispersion interactions, the hydrophobic effect (Kauzmann, 1959), hydrogen bonds (Baker & Hubbard, 1984;Fersht et al, 1985;Yang et al, 1992), salt bridges (Fersht, 1971(Fersht, , 1972Marqusee & Baldwin, 1987;Anderson et al, 1990;Horovitz et al, 1990;Serrano et al, 1990;Dao-pin et al, 1991;Merutka & Stellwagen, 1991;Lyu et al, 1992;HuyghuesDespointes et al, 1993;Hendsch & Tidor, 1994;Waldburger et al, 1995;Wimley et al, 1996) and local backbone and side-chain entropy (Page & Jencks, 1971;Matthews et al, 1987;Straatsma & McCammon, 1989;Tobias et al, 1989;Pickett & Sternberg, 1993) to include more aggregate and delocalized effects. Examples range from the medium-ranged, such as helix-capping interactions (Hol et al, 1978;Sheridan et al, 1982;Hol, 1985;Shoemaker et al, 1987;Nicholson et al, 1988Nicholson et al, , 1991Presta & Rose, 1988;Richardson & Richardson, 1988;Sali et al, 1988;Fairman et al, 1989;Serrano & Fersht, 1989;Lyu et al, 1993;Hendsch & Tidor, 1994;Tidor, 1994;PrCvost, 1996) to the global, such as overall vibrational entropy …”

“…Recent theoretical and experimental evidence has suggested that an individual salt bridge may be less stabilizing than a structurally equivalent hydrophobic interaction (Hendsch & Tidor, 1994;Waldburger et al, 1995;Wimley et al, 1996). That is, salt bridges in proteins are generally destabilizing relative to replacement with hydrophobic groups of the same size and shape.…”

“…Indeed, some experimental evidence for an increase in degeneracy upon replacement of charged or polar interactions with hydrophobic ones has been seen in the coiled coils (Harbury et al, 1993;Lumb & Kim, 1995). In experiments using the Arc repressor, no such increase in structural degeneracy was seen (Waldburger et al, 1995). This could be due to a larger network of cooperative interactions defining the ground-state structure in the Arc repressor as compared with the coiled coils.…”

Putative Interhelix Ion Pairs Involved in the Stability of Myoglobin

“…Similarly, an Asp-Arg interaction makes a major contribution to the stability of barnase (32). On the other hand, Arc repressor contains a complex ion pair interaction involving three partially buried charged side chains and replacement of all three charged residues by nonpolar residues increases its stability (33). Side chain ion pairs, at specific spacings of i, i + 4, stabilize peptide helices by modest but measurable amounts (3,4).…”