2.1. Data Sets. Bacteriophage T4 lysozyme promotes the release of phage particle from the wall of a cell by breaking down peptidoglycan, allowing for the injection of genomic DNA into the host Escherichia coli cell. The class of enzymes considered here have apolar and polar binding sites and are known as L99A and L99A/M102Q, respectively. For these systems, calculation of protein-ligand interaction energies has proven challenging for classical molecular dynamics methods. Benchmark data sets of crystal structures and binding energies for both sites, with a variety of non-covalent ligands, were introduced in a recent review. These examples having binding energies within a narrow range from 4.0-6.7 kcal/mol, with estimated uncertainties that are < 0.2 kcal/mol. We selected two representative systems from the L99A data set, with protein databank (PDB) codes 181L and 4W54. The L99A/M102Q data set introduces a point mutation at one side of the binding site, replacing methionine residue 102 with the polar side chain of glutamine to serve as a hydrogen-bond acceptor. From this data set we selected representative systems 1LI2 and 3HUA. The ligands for these four T4-lysozyme complexes are benzene (for 181L), ethylbenzene (for 4W54), phenol (for 1LI2), and indole (for 3HUA); see Fig. .