In Figure , we show the distributions of the λ-coordinate in five constant pH MD replicas of the ADA system with the original CHARMM36m force field parameters. Distributions of the λ-coordinates in the other systems (AEA, AKA AHA, and AAA 1 and AAA 2 ) are shown in Supporting Information (SI, Figures ). The dissimilarity between the λ-distributions in the replicas (maximum KSS between replicas of 0.29, 0.11, 0.04, and 0.095 for ADA, AEA, AHA, and AAA 1 , respectively) indicates a lack of convergence. In addition, the distributions of the dihedral angles, shown in Figure , c, and d, are also not identical for all replicas. Because there is no barrier from the biasing potential for the λ-coordinate, we conclude that the lack of convergence in λ is due to insufficient sampling of the dihedral degrees of freedom.