Beyond inhibition: reversing a bacterial multidrug efflux pump

Katherine Henzler-Wildman

Univ. of Wisconsin-Madison

Thursday, November 21, 2019 - 11:30am

Lederle Graduate Research Tower 1634

Reception and light refreshments at 11:00 a.m.

EmrE is small multidrug transporter found in the inner membrane of E. coli. We are interested in how this very small transporter harnesses the proton motive force to actively pump polyaromatic cations out of the bacteria, thus conferring resistance to a wide range of toxic compounds. EmrE is an asymmetric, antiparallel homodimer with a single substrate binding site defined by a pair of glutamates in the dimer interface. Like all active transporters, EmrE must change conformation to allow alternating access of the binding site to either side of the membrane in order to move bound substrate across the membrane from one aqueous compartment to another. We have used NMR to directly monitor proton binding and alternating access in EmrE. Our NMR data revealed additional states and conformational transitions that must be included in the transport model. Our new free exchange model predicts that EmrE should be able to perform antiport with multiple drug/proton stoichiometries, as well as uniport and even drug/proton symport. Mathematical modeling demonstrates that drug/proton antiport remains the dominant transport pathway near physiological conditions. Liposomal flux assays confirm that EmrE is not a tightly coupled antiporter and can perform drug/proton transport with different stoichiometries under different environmental conditions. Previous studies of multidrug recognition in EmrE demonstrated that the rate of alternating access and drug efflux depend on the identity of the transported substrate. We are now using in vivo assays to test whether the identity of the transported substrate can also determine the direction of proton-coupled transport. These results change our understanding of how this promiscuous transporter functions and suggest new mechanisms for targeting antibiotic resistance due to drug efflux. 

Speaker Link:
Faculty Host: Lynmarie Thompson
Event Contact:
Laura Sedberry
lsedberry@chem.umass.edu
(413) 545-2585