In general, the sign and magnitude of the RMSD difference between the bound and unbound states ∆G RM SD bound + ∆G RM SD unbound indicates whether the re-arrangement of the 'receptor' helices is favourable or not. A positive RMSD difference indicates that the 'receptor' helices are allowed to re-arrange more in the unbound state where a helix has been removed, compared to the bound state. On the contrary, a negative RMSD difference indicates that the 'receptor' helices are allowed to re-arrange more in the bound state compared to the unbound state. This can explain the RMSD differences observed in the three dissociation steps of CC-Tet. In the tetramer step, any re-arrangement of the 'receptor' helices is limited in the bound state due to the presence of all four helices in a 'closed' conformation, and the RMSD difference is therefore highly positive. In the trimer step, the absence of one helix and the 'open' conformation of the 'receptor' confers more re-arrangement freedom to the 'receptor' helices in the bound state; the RMSD difference in this case is still positive, albeit less positive compared to the tetramer step. In the dimer step, the RMSD difference is negative, indicating that the intermediate dimer can re-arrange more in the bound state than in the unbound state. This is an interesting observation, as one might expect a single α-helix in the bulk to have more conformational freedom than a dimeric complex, especially one that adopts an 'open' conformation; however, the ϕ, ψ torsional restraints applied throughout the free energy calculations limit the conformations available to the monomeric α-helix in the bulk. In general, all intermediate dimer dissociation steps have small absolute ∆G RM SD bound + ∆G RM SD unbound values, indicating that there are small differences in the 'receptor' re-organisation potential between the bound and unbound states.