Absolute Binding Free Energy Calculations for Buried Water Molecules

Water often plays a key role in mediating protein-ligand interactions. Understanding contributions from active-site water molecules to binding thermodynamics of a ligand is important in predicting binding free energies for ligand optimization. In this work, we tested a nonequilibrium switching method for absolute binding free energy calculations on water molecules in binding sites. We studied five protein targets with different ligands for which the binding free energies of selected buried water sites had been studied by previous work using a different simulation technique. Our calculated water binding free energies agree well with literature values. We also discuss the lessons we learned about identified issues that affected our calculations and ways to address them. This work fits with our larger focus on how to do accurate ligand binding free energy calculations when water rearrangements are very slow, such as rearrangements due to ligand modification (as in relative free energy calculations) or ligand binding (as in absolute free energy calculations). The method studied in this work can potentially be used to account for limited water sampling via providing endpoint corrections to free energy calculations using our calculated binding free energy of water.