Improving pKa prediction accuracy with reparametrized force fields and free energy calculations

Given the growing interest in designing targeted covalent inhibitors, methods for rapidly and accurately probing pKas—and, by extension, the reactivities—of target cysteines is highly desirable. Complementary to cysteine, histidine is similarly relevant due to its frequent presence in protein active sites and its unique ability to exist in two tautomeric states. Here we demonstrate that non-equilibrium free energy calculations can accurately determine the pKa values of both residues, often outperforming conventional predictors. Importantly, we find that: 1) increasing the van der Waals radii of cysteine's sulfur atom; 2) altering the backbone charges of histidine; and 3) downscaling the side chain partial charges of both residues, can all significantly improve pKa prediction accuracy. Using the modified CHARMM36m force field, we achieve an average unsigned error of 1.61±0.21 pK for cysteine and 0.71±0.16 pK for histidine, while using the modified Amber14SB force field yields errors of 2.37±0.29 pK for cysteine and 0.50±0.10 pK for histidine.