Cryo-EM structure of the EspA filament from enteropathogenic Escherichia coli: Revealing the mechanism of effector translocation in the T3SS

Abstract

The type III secretion system (T3SS) is a virulence mechanism employed by Gram-negative pathogens. The T3SS forms a proteinaceous channel that projects a needle into the extracellular medium where it interacts with the host cell to deliver virulence factors. Enteropathogenic Escherichia coli (EPEC) is unique in adopting a needle extension to the T3SS-a filament formed by EspA-which is absolutely required for efficient colonization of the gut. Here, we describe the cryoelectron microscopy structure of native EspA filaments from EPEC at 3.6-Å resolution. Within the filament, positively charged residues adjacent to a hydrophobic groove line the lumen of the filament in a spiral manner, suggesting a mechanism of substrate translocation mediated via electrostatics. Using structure-guided mutagenesis, in vivo studies corroborate the role of these residues in secretion and translocation function. The high-resolution structure of the EspA filament could aid in structure-guided drug design of antivirulence therapeutics.

Keywords: EPEC virulence; T3SS; antibiotic resistance; cryoelectron microscopy; vaccine targets.