Molecular Dynamics Investigation of MFS Efflux Pump MdfA Reveals an Intermediate State between Its Inward and Outward Conformations

Multidrug resistance poses a major challenge to antibiotic therapy. A principal cause of antibiotic resistance is through active export by efflux pumps embedded in the bacterial membrane. Major facilitator superfamily (MFS) efflux pumps constitute a major group of transporters, which are often related to quinolone resistance in clinical settings. Although a rocker-switch model is proposed for description of their conformational transitions, detailed changes in this process remain poorly understood. Here we used MdfA from E. coli as a representative MFS efflux pump to investigate factors that can affect its conformational transition in silico. Molecular dynamics (MD) simulations of MdfA’s inward and outward conformations revealed an intermediate state between these two conformations. By comparison of the subtle differences between the intermediate state and the average state, we indicated that conformational transition from outward to inward was initiated by protonation of the periplasmic side. Subsequently, hydrophilic interaction of the periplasmic side with water was promoted and the regional structure of helix 1 was altered to favor this process. As the hydrophobic interaction between MdfA and membrane was also increased, energy was concentrated and stored for the opposite transition. In parallel, salt bridges at the cytoplasmic side were altered to lower probabilities to facilitate the entrance of substrate. In summary, we described the total and local changes during MdfA’s conformational transition, providing insights for the development of potential inhibitors.