Free Energy Calculations of Mutations Involving a Tightly Bound Water Molecule and Ligand Substitutions in a Ligand-Protein Complex

The accurate calculation of the free energy of interaction of protein-water-ligand systems has an important role in molecular recognition and drug design that is often not fully considered. We report free energy thermodynamic integration calculations used to evaluate the effects of inclusion, neglect, and targeting and removal (i.e., systematic substitution by ligand functional groups) of an important, tightly bound, water molecule in the SH3 domain of Abl tyrosine kinase. The effects of this water molecule on the free energies of interaction of several Abl-SH3 domain-ligand systems reveal that there is an unfavourable free energy change associated with its removal into the bulk solvent. Only three substitutions by an additional functional group (out of methyl, ethyl, hydroxyl, amino, and amide groups) in the phenyl ring of a tyrosine in the peptide ligand resulted in a favourable change in the free energy of binding upon replacement of the ordered water molecule. This computational approach provides a direct route to the systematic and rigorous prediction of the thermodynamic influence of ordered, structural water molecules on ligand modification and optimization in drug design by calculating free energy changes in protein-water-ligand systems.