Molecular insights into ATP-mediated NBD dimerization in an ABC transporter.

CmABCB1 is a Cyanidioschyzon merolae homolog of human ABCB1, which is a member of the ATP-binding cassette (ABC) transporter superfamily responsible for the efflux of a wide range of substrates from cells. The two major conformations of CmABCB1 are the inward-facing conformation that binds the substrate to be transported, and the outward-facing conformation that represents the state post the transport of the substrate. In this study, we have performed a 1000 ns all-atom MD simulation of CmABCB1 with and without ATP to understand how ATP binding influences the dynamics and conformation of the protein. Additionally, we have also performed two distinct methods of umbrella sampling (US) simulations to determine the free energy of binding of the nucleotide-binding domains (NBDs) both in the presence and absence of ATP. Our MD simulations reveal significant structural differences of the transporter depending on whether ATP is present or absent at the NBDs. Only when ATP was present at the NBDs, we discovered a specific salt-bridge interaction between the coupling helix (CH) and the nucleotide-binding domain (NBD), which we believe could play a potential role in substrate transport and the accompanying conformational change to the outward-facing state. We also observed a significant loss in the NBD-NBD interactions in the absence of ATP. Our umbrella sampling simulations showed that ATP binding stabilizes the NBD dimer by about ∼25 kJ/mol. Overall, our findings provide valuable insights into the conformational changes of CmABCB1 and the role of ATP in the transport cycle of ABC transporters.