Free energies of protein decoys provide insight into determinants of protein stability

We have calculated the stability of decoy structures of several proteins (f rom the CASP3 models and the Park and Levitt decoy set) relative to the nat ive structures. The calculations were performed with the force field-consis tent ES/IS method, in which an implicit solvent (IS) model is used to calcu late the average solvation free energy for snapshots from explicit simulati ons (ESs). The conformational free energy is obtained by adding the interna l energy of the solute from the ESs and an entropic term estimated from the covariance positional fluctuation matrix. The set of atomic Born radii and the cavity-surface free energy coefficient used in the implicit model has been optimized to be consistent with the all-atom force field used in the E Ss (cedar/gromos with simple point charge (SPC) water model). The decoys ar e found to have a consistently higher free energy than that of the native s tructure; the gap between the native structure and the best decoy varies be tween 10 and 15 kcal/mole, on the order of the free energy difference that typically separates the native state of a protein from the unfolded state. The correlation between the free energy and the extent to which the decoy s tructures differ from the native (as root mean square deviation) is very we ak; hence, the free energy is not an accurate measure for ranking the struc turally most native-like structures from among a set of models. Analysis of the energy components shows that stability is attained as a result of thre e major driving forces: (1) minimum size of the protein-water surface inter face: (2) minimum total electrostatic energy, which includes solvent polari zation; and (3) minimum protein packing energy. The detailed fit required t o optimize the last term may underlie difficulties encountered in recoverin g the native fold from an approximate decoy or model structure.