Contribution of polar groups in the interior of a protein to the conformational stability

It has been generally believed that polar residues are usually located on t he surface of protein structures, However, there are many polar groups in t he interior of the structures in reality. To evaluate the contribution of s uch buried polar groups to the conformational stability of a protein, nonpo lar to polar mutations (L8T, A9S, A32S, I56T, I59T, I59S, A92S, V93T, A96S, V99T, and V100T) in the interior of a human lysozyme were examined. The th ermodynamic parameters for denaturation were determined using a differentia l scanning calorimeter, and the crystal structures were analyzed by X-ray c rystallography. If a polar group had a heavy energy cost to be buried, a mu tant protein would be remarkably destabilized. However, the stability (Delt aG) of the Ala to Ser and Val to Thr mutant human lysozymes was comparable to that of the wild-type protein, suggesting a low-energy penalty of buried polar groups. The structural analysis showed that all polar side chains in troduced in the mutant proteins were able to find their hydrogen bond partn ers, which are ubiquitous in protein structures. The empirical structure-ba sed calculation of stability change (Delta DeltaG) [Takano et al. (1999) Bi ochemistry 38, 12698-12708] revealed that the mutant proteins decreased the hydrophobic effect contributing to the stability (DeltaG(HP)), but this de stabilization was recovered by the hydrogen bonds newly introduced. The pre sent study shows the favorable contribution of polar groups with hydrogen b onds in the interior of protein molecules to the conformational stability.