Release of Entropic Spring Reveals Conformational Coupling Mechanism in the ABC Transporter BtuCD-F

Abstract

Substrate translocation by ATP-binding cassette (ABC) transporters involves coupling of ATP binding and hydrolysis in the nucleotide-binding domains (NBDs) to conformational changes in the transmembrane domains. We used molecular dynamics simulations to investigate the atomic-level mechanism of conformational coupling in the ABC transporter BtuCD-F, which imports vitamin B12 across the inner membrane of Escherichia coli. Our simulations show how an engineered disulfide bond across the NBD dimer interface reduces conformational fluctuations and hence configurational entropy. As a result, the disulfide bond is under substantial mechanical stress. Releasing this entropic spring, as is the case in the wild-type transporter, combined with analyzing the pairwise forces between individual residues, unravels the coupling mechanism. The identified pathways along which force is propagated from the NBDs via the coupling helix to the transmembrane domains are composed of highly conserved residues, underlining their functional relevance. This study not only reveals the details of conformational coupling in BtuCD-F, it also provides a promising approach to other long-range conformational couplings, e.g., in ABC exporters or other ATP-driven molecular machines.

Molecular Mechanism of ATP Hydrolysis in an ABC Transporter

“…We initiated our QM/MM simulations from a representative hydrolysis-competent snapshot taken from our previous force field MD simulations. 54 …”

“… 43 , 44 In this conformation, the occupancy of the binding pocket with water molecules is significantly enhanced. 54 This increased hydration of the binding pocket could be one of the initial steps on the way to the release of the reaction products, as water molecules can replace hydrogen bonds of ADP and P i with the protein.…”

“…The QM/MM MD simulations were started from pre-equilibrated snapshots taken from our recent force field MD study, 54 and we hence refer to this previous study for the details of the simulation setup. In brief, the complete BtuCD-F complex was simulated in a fully solvated palmitoyloleoylphosphatidylcholine (POPC) lipid bilayer.…”

Comparison of force fields for Alzheimer's A: A case study for intrinsically disordered proteins

“…At every exchange interval, 32768 ( , where is the number of replicas) random exchanges were attempted as suggested by Chodera and Shirts . Virtual sites were used for all protein hydrogens which permitted a time step of 4 fs to be used, a strategy that has been successfully employed before . Protein bonds were constrained using the P‐Lincs algorithm with an expansion order of 6 and water molecules were constrained with the Settle algorithm .…”

“…61 Virtual sites were used for all protein hydrogens 62 which permitted a time step of 4 fs to be used, a strategy that has been successfully employed before. [63][64][65][66] Protein bonds were constrained using the P-Lincs algorithm 67 with an expansion order of 6 and water molecules were constrained with the Settle algorithm. 68 Electrostatic interactions were calculated with the particle mesh Ewald method 69 with a Fourier spacing of 0.12 nm and a short-range cutoff of 1.2 nm.…”