Josh V Vermaas, Susan B. Rempe, and Emad Tajkhorshid.
Electrostatic lock in the transport cycle of the multi-drug
resistance transporter EmrE.
Proceedings of the National Academy of Sciences, USA,
115:E7502-E7511, 2018.
(PMC: PMC6094130)
VERM2018-ET
EmrE is a small, homodimeric membrane transporter that exploits the
established electrochemical proton gradient across the Escherichia coli inner
membrane to export toxic polyaromatic cations, prototypical of the wider
small-multidrug resistance transporter family. While prior studies have
established many fundamental aspects of the specificity and rate of
substrate transport in EmrE, low resolution of available structures has
hampered identification of the transport coupling mechanism. Here we
present a complete, refined atomic structure of EmrE optimized against
available cryo-electron microscopy (cryo-EM) data to delineate the critical
interactions by which EmrE regulates its conformation during the transport
process. With the model, we conduct molecular dynamics simulations of the
transporter in explicit membranes to probe EmrE dynamics under different
substrate loading and conformational states, representing different
intermediates in the transport cycle. The refined model is stable under
extended simulation. The water dynamics in simulation indicate that the
hydrogen-bonding networks around a pair of solvent-exposed glutamate
residues (E14) depend on the loading state of EmrE. One specific hydrogen
bond from a tyrosine (Y60) on one monomer to a glutamate (E14) on the
opposite monomer is especially critical, as it locks the protein conformation
when the glutamate is deprotonated. The hydrogen bond provided by Y60
lowers the pKa of one glutamate relative to the other, suggesting both
glutamates should be protonated for the hydrogen bond to break and a
substrate-free transition to take place. These findings establish the
molecular mechanism for the coupling between proton transfer reactions
and protein conformation in this proton-coupled secondary transporter.
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