Characterizing Transition Pathways in the Transport Cycle of ABC Transporter MsbA

Abstract

MsbA is a member of the ABC (ATP-binding cassette) family transporters that uses energy from ATP hydrolysis to transport various substrates across cellular membrane. The transport cycle of MsbA has been described by a simple “Alternating Access” mechanism in which the transporter changes its conformation between an outward-facing (OF) and an inward-facing (IF) state while coupling the translocation of substrates. Here, to investigate the detailed transport cycle of MsbA at atomistic resolution, we have performed molecular dynamics (MD) simulations using a combination of several computational methods. Starting from the recently solved X-ray crystal structure of MsbA in OF state, we first generate high resolution all-atom models for the two IF states (IF open and IF closed) by using targeted MD simulations. We then define two collective variables to describe the relative motion of the two transmembrane domains (TMDs) and nucleotide binding domains (NBDs), respectively. Steered MD simulations along the collective variables have been performed to induce the conformational changes between the three states (OF, IF open and IF closed) of MsbA. Assessing the energetics associated with the induced transitions, approximated by calculating the non-equilibrium work involved in going from one state to another suggest that the OF-to-IF conformational transition follows two steps: the two TMDs close first, and then the two NBDs open, as opposed to the pathway observed from initial targeted MD simulations. Taken together, these results not only provide a better understanding of the functionality of ABC transporters, but also help define a general mechanism for membrane transport process.