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Exploring the Mechanism of the Ion-Coupled Transporter EmrE
Maureen Leninger, Anindita Gayen, Nathaniel J. Traaseth
Department of Chemistry, New York University, New York, NY 10012
Molecular transport of toxic molecules such as antiseptics and antibiotics by efflux pumps is one of the primary
1,2
mechanisms that enable bacteria to gain multidrug resistance. While it is well known that these efflux pumps use the
proton motive force across the Escherichia coli inner membrane to efflux drugs against the concentration gradient, the
molecular details for this transport process is still unknown. EmrE is a member of the small multidrug resistance family
that we used to elucidate the mechanism on an atomic scale. A combination of solution and solid-state NMR along with
biochemical resistance assays revealed that the pKa of Glu-14, a conserved glutamate residue in the binding pocket,
controls the resistance phenotype of EmrE by altering the dynamics of the transporter. Deprotonation of Glu-14 reduced
the rate of conformational exchange by 6-fold. The difference in rates between these protonated and deprotonated EmrE
at Glu-14 biases EmrE in an inward-open conformation positioned towards the cytoplasm to bind and transport toxic
compounds. Interestingly although the deprotonated transporter has a reduced exchange rate it is not completely halted.
These findings are important in understanding how EmrE is responsible for sensitizing E. coli to toxic compounds in
conditions where the pH gradient is exclusively responsible for driving transport. We have also been able to identify
certain residues in the loop between helix 2 and 3 that are involved in the stabilization of the transporter in the
deprotonated conformation. When these residues are mutated the rate of conformational exchange is affected along with
the resistance phenotype. The combination of NMR spectroscopy and biochemical resistance assays provided insight into
the ion-coupled transport mechanism of EmrE.
References
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bacteria. FEMS Microbiol Rev 36, 340-63 (2012).
Allen, H.K.D., Justin; Wan, Helena Huimi; Cloud-Hansen, Karen A; Davies, Julian; Handelsman, Jo. Call of the
wild: antibiotic resistance genes in natural environments. Nature Reviews Microbiology, 251-259 (2010).