Additionally, as a distance metric, RMSD does not account for the potential size of states. As an example, we observe that frames in the top left corner of the half-moon shaped well and frames in the right side of the oval-shaped well are similarly distant to the 4w59 structure based on RMSD (Figure ). However, we instead find with our kinetics-based definition that the entirety of the half-moon shaped well describes the open state (Figure ). While the top and bottom portions of the half-moon shaped well can be further separated into kinetically distinct states (Figure ), this separation is entirely along the second tIC, which does not primarily correspond to opening of the F-helix.