Electrostatic lock in the transport cycle of the multidrug resistance transporter EmrE

Abstract

SignificanceEmrE is a small membrane transporter found in Escherichia coli that exports drug-like molecules from the cell, contributing to antibiotic resistance. In EmrE, as well as in the wider small-multidrug resistance transporter family, a specific anionic amino acid (E14) has been implicated in governing the conformational changes that export drugs. However, due to sparse structural information, the exact interactions remain unidentified. Through interactive molecular dynamics to incorporate existing cryo-electron microscopy data, we create a fully refined atomic model of EmrE. We then embed this model in a lipid bilayer and evaluate the interactions within EmrE under different loading states. We find that E14 makes specific hydrogen bonds to neighboring residues, coupling observed experimental phenomena to interactions at the atomic scale.

Zooming in on a small multidrug transporter reveals details of asymmetric protonation

“…Having verified that the refined model is stable in the five loading states, Vermaas et al (10) examined the hydration of the interior cavity (lumen) of EmrE. As expected from the inwardfacing starting structure, A − B − , A H B − , and A − B H displayed water channels that connect the lumen to the cytoplasmic side.…”

“…In PNAS, Vermaas et al (10) report a computational study that combines a battery of state-of-theart modeling and simulation tools to shed new light on the perplexing drug-proton antiport mechanism of EmrE at the atomic level. To enable the study, the authors first constructed a complete atomic model based on the 3.8-Å resolution, C α -only crystal structure of EmrE in complex with tetraphenylphosphonium (TPP + ) (4).…”

“…The atomic model was then refined to the so-called fully atomic refined model (faRM) using molecular dynamics (MD) flexible fitting to the cryoelectron microscopy density map of EmrE at 7.5-Å resolution (11). The faRM was validated by running 500-ns all-atom MD simulations of EmrE embedded in a fully explicit dimyristoylphosphatidylcholine (DMPC) bilayer, whereby EmrE was in one of the five substrate/proton loading states: A − B − , A − B − TPP + , A H B H , A H B − , and A − B H , which are denoted apo, TPP + , A + B + , A + , and B + , respectively, in the paper by Vermaas et al (10). While A − B − , A − B − TPP + , and A H B H are the only possible states in the two-proton model, A H B − and A − B H are also possible in the one-proton model ( Fig.…”

“…1). Note that the singly protonated drug-bound state, A H B − TPP + or A − B H TPP + , was not studied by Vermaas et al (10).…”

“…To circumvent the limitation in the simulation length, which makes the direct simulation of the inward-outward conformational transition prohibitive, Vermaas et al (10) applied steered MD to the different loading states, using the rmsd between the initial inward-and target outward-facing structures as the biasing force. The target structure was built by taking advantage of the symmetry of the antisymmetric homodimer.…”