The C terminus of the bacterial multidrug transporter EmrE couples drug binding to proton release

Nathan E. Thomas, Chao Wu, Emma A. Morrison, Anne E. Robinson, Josephine P. Werner, Katherine A. Henzler-Wildman

Research output: Contribution to journalReview articlepeer-review

9 Scopus citations

Abstract

Ion-coupled transporters must regulate access of ions and substrates into and out of the binding site to actively transport substrates and minimize dissipative leak of ions. Within the single-site alternating access model, competitive substrate binding forms the foundation of ion-coupled antiport. Strict competition between substrates leads to stoichiometric antiport without slippage. However, recent NMR studies of the bacterial multidrug transporter EmrE have demonstrated that this multidrug transporter can simultaneously bind drug and proton, which will affect the transport stoichiometry and efficiency of coupled antiport. Here, we investigated the nature of substrate competition in EmrE using multiple methods to measure proton release upon the addition of saturating concentrations of drug as a function of pH. The resulting proton-release profile confirmed simultaneous binding of drug and proton, but suggested that a residue outside EmrE’s Glu-14 binding site may release protons upon drug binding. Using NMR-monitored pH titrations, we trace this drug-induced deprotonation event to His-110, EmrE’s C-terminal residue. Further NMR experiments disclosed that the C-terminal tail is strongly coupled to EmrE’s drug-binding domain. Consideration of our results alongside those from previous studies of EmrE suggests that this conserved tail participates in secondary gating of EmrE-mediated proton/drug transport, occluding the binding pocket of fully protonated EmrE in the absence of drug to prevent dissipative proton transport.

Original languageEnglish
Pages (from-to)19137-19147
Number of pages11
JournalJournal of Biological Chemistry
Volume293
Issue number49
DOIs
StatePublished - Dec 7 2018

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