As indicated in Section 3.1, we sought insight into the computed relative binding enthalpies by isolating the mean energies associated with parts of the overall system, notably the ligand and a set of residues that form the binding site. To generate ligand-only trajectories, we used cpptraj to delete the protein and all solvent molecules, including water, HEPES buffer, and ions, from each trajectory. We similarly generated trajectories containing only 13 binding site residues (Arg13, Arg32, Ser34, Glu35, Ser36, Ser42, Val51, Gln52, His53, Phe54, Lys55, Leu66, Trp67), with and without the bound ligand; the rest of the protein and all solvent molecules were stripped. The potential energies of the decomposed systems were then evaluated by specifying imin=5 and max-cyc=1 in the sander program to read in the trajectories and calculate a single-point energy at each frame. PME was disabled by using ntb=0 so that no periodicity was applied and long-ranged interactions were accounted for instead by increasing the nonbonded cutoff to 100.0 Å.