We additionally ran Boresch-restrained metadynamics simulations of a structure of CC-Di designed in ISAMBARD, to compare to the simulations of the X-ray structure. The same anchor points and force constants for the Boresch restraints were used. Equal binding free energies were obtained for the two structures within error (Table ), showing that our protocol performs equally well on experimental and designed structures, provided the designed structures are reliable. This comparison is especially significant for CC-Di, since its asymmetric unit in the X-ray structure does not contain the dimer interface. For the remainder of our simulations, we used experimental structures where they were available, and designed any alternative states with ISAMBARD.