Molecular dynamics simulations of the hyperthermophilic protein sac7d from Sulfolobus acidocaldarius: contribution of salt bridges to thermostability.

Hyperthermophilic proteins often possess an increased number of surface salt bridges compared with their mesophilic homologues. However, salt bridges are generally thought to be of minor importance in protein stability at room temperature. In an effort to understand why this may no longer be true at elevated temperatures, we performed molecular dynamics simulations of the hyperthermophilic protein Sac7d at 300 K, 360 K, and 550 K. The three trajectories are stable on the nanosecond timescale, as evidenced by the analysis of several time-resolved properties. The simulations at 300 K and (to a lesser extent) 360 K are also compatible with nuclear Overhauser effect-derived distances. Raising the temperature from 300 K to 360 K results in a less favourable protein-solvent interaction energy, and a more favourable intraprotein interaction energy. Both effects are almost exclusively electrostatic in nature and dominated by contributions due to charged side-chains. The reduced solvation is due to a loss of spatial and orientational structure of water around charged side-chains, which is a consequence of the increased thermal motion in the solvent. The favourable change in the intraprotein Coulombic interaction energy is essentially due to the tightening of salt bridges. Assuming that charged side-chains are on average more distant from one another in the unfolded state than in the folded state, it follows that salt bridges may contribute to protein stability at elevated temperatures because (i) the solvation free energy of charged side-chains is more adversely affected in the unfolded state than in the folded state by an increase in temperature, and (ii) due to the tightening of salt bridges, unfolding implies a larger unfavourable increase in the intraprotein Coulombic energy at higher temperature. Possible causes for the unexpected stability of the protein at 550 K are also discussed. Copyright 1999 Academic Press.