Binding of buried structural water increases the flexibility of proteins

Water deeply buried in proteins is considered to be an integral part of the folded structure. Such structural water molecules make strong H bonds with polar groups of the surrounding protein and therefore are believed to tighten the protein matrix. Surprisingly, our computational analysis of the binding of a buried water molecule to bovine pancreatic trypsin inhibitor shows that the protein actually becomes more flexible, as revealed by an increase in the vibrational entropy. We find that this effect must be common in proteins, because the large entropic cost of immobilizing a single water molecule [−TΔS = 20.6 kcal/mol (1 kcal = 4.18 kJ) for the lost translational and rotational degrees of freedom] can only be partly compensated by water–protein interactions, even when they are nearly perfect, as in the case of bovine pancreatic trypsin inhibitor (ΔE = −19.8 kcal/mol), leaving no room for a further decrease in entropy from protein tightening. This study illustrates the importance of considering changes in protein flexibility (which in this case favor binding by 3.5 kcal/mol) for the prediction of ligand binding affinities.