We tested SepTop on a larger dataset, 16 ligands binding to MALT1. We also tested SepTop on a larger dataset, examining a series of 16 ligands binding to MALT1, and obtained good correlation and error statistics. We created a ligand map in which the 16 ligands were connected with 30 edges. To setup the ligand map, we chose one of the most potent ligands as the reference ligand and connected all other ligand to that reference ligand through edges, creating a star map. We then added additional edges between ligands to create ligand cycles. Absolute binding free energies were obtained from calculated ∆∆G values and experimental binding free energies using the arsenic code. Since the experimental binding affinities for 6 of the compounds were outside the assay limit, we excluded those ligands from the correlation and error statistics. Correlating the calculated binding free energy of the 10 remaining ligands with the experimental values resulted in an RM SE = 1.06 and R 2 = 0.85. Additional statistics can be found in Figure . The full set of calculated and experimental values can be found in the SI. Figure . Different choices made in ER alpha preparation. (a) Two loops were not resolved in the crystal structure and missing residues were modelled in some studies, while in other studies the residues bordering the missing loop were capped and restraints used to keep the protein from drifting apart. (b) Different software for the assignment of protein side chain protonation states resulted in differences in protonation state of some histidine residues. One of those residues is located in the binding and the selected protonation state impacted the ∆∆G estimate, at least in our study.