the trajectories reveals that the larger RMSDs with HG-optimized radii are due to increased flexibility in some alpha helices. This makes sense, as the HG-optimized shrinks the GB cavity radii for hydrogen and nitrogen atoms, resulting in weaker backbone hydrogen bonds along the helix. Superposed representative structures of the four proteins, comparing their conformations in TIP3P and OBC2(HG-optimized) solvent, are shown in Figure . Despite the increased flexibility in some protein regions, all four proteins maintained their tertiary structure with the HG-optimized radii. We also note that the ligands were stable in their binding pockets during the ABFE calculations. and.