One such slow motion which may have had an impact here is the re-arrangement of one of the binding site loops in the region of residues ASN106 through to SER113. As shown in Fig. , the helicity of this loop region changes between the initial models used for ligands 3-20, 24, In this study six elaboration decisions were outlined, of these three would have been clearly supported by our ABFE results (SI Table ). Of the three failed decisions only one (decision 2), would have been fully miscalculated. For the other two, one involved small changes in experimental affinity (decision 5) with ΔΔG values ~ 0.6-0.8 kcal/mol. This likely still falls within the accuracy of ITC measurement, but is too small a change to be properly captured by ABFE given the size of sampling errors. The other elaboration decision clearly identified improvements in affinity but failed to identify ligand 24 as a better binder than ligands 25 and 26 (decision 4).