Conformational dynamics and putative substrate extrusion pathways of the N-glycosylated outer membrane factor CmeC from Campylobacter jejuni.

Abstract

The outer membrane factor CmeC of the efflux machinery CmeABC plays an important role in conferring antibiotic and bile resistance to Campylobacter jejuni. Curiously, the protein is N-glycosylated, with the glycans playing a key role in the effective function of this system. In this work we have employed atomistic equilibrium molecular dynamics simulations of CmeC in a representative model of the C. jejuni outer membrane to characterise the dynamics of the protein and its associated glycans. We show that the glycans are more conformationally labile than had previously been thought. The extracellular loops of CmeC visit the open and closed states freely suggesting the absence of a gating mechanism on this side, while the narrow periplasmic entrance remains tightly closed, regulated via coordination to solvated cations. We identify several cation binding sites on the interior surface of the protein. Additionally, we used steered molecular dynamics simulations to elucidate translocation pathways for a bile acid and a macrolide antibiotic. These, and additional equilibrium simulations suggest that the anionic bile acid utilises multivalent cations to climb the ladder of acidic residues that line the interior surface of the protein.