title: Specificity determinants in small multidrug transporters.
abstract: Multiple-antibiotic resistance has become a major global public health concern, and to overcome this problem, it is necessary to understand the resistance mechanisms that allow survival of the microorganisms at the molecular level. One mechanism responsible for such resistance involves active removal of the antibiotic from the pathogen cell by MDTs (multidrug transporters). A prominent MDT feature is their high polyspecificity allowing for a single transporter to confer resistance against a range of drugs. Here we present the molecular mechanism underlying substrate recognition in EmrE, a small MDT from Escherichia coli. EmrE is known to have a substrate preference for aromatic, cationic compounds, such as methyl viologen (MV(2+)). In this work, we use a combined bioinformatic and biochemical approach to identify one of the major molecular determinants involved in MV(2+) transport and resistance. Replacement of an Ala residue with Ser in weakly resistant SMRs from Bacillus pertussis and Mycobacterium tuberculosis enables them to provide robust resistance to MV(2+) and to transport MV(2+) and has negligible effects on the interaction with other substrates. This shows that the residue identified herein is uniquely positioned in the binding site so as to be exclusively involved in the mediating of MV(2+) transport and resistance, both in EmrE and in other homologues. This work provides clues toward uncovering how specificity is achieved within the binding pocket of a polyspecific transporter that may open new possibilities as to how these transporters can be manipulated to bind a designed set of drugs.
authors.affiliation: Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, Hebrew University of Jerusalem, 91904 Jerusalem, Israel.
authors.forename: Shlomo
authors.initials: S
authors.lastname: Brill
authors.affiliation: Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, Hebrew University of Jerusalem, 91904 Jerusalem, Israel.
authors.forename: Ofir
authors.initials: O
authors.lastname: Sade-Falk
authors.affiliation: Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, Hebrew University of Jerusalem, 91904 Jerusalem, Israel.
authors.forename: Yael
authors.initials: Y
authors.lastname: Elbaz-Alon
authors.affiliation: Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, Hebrew University of Jerusalem, 91904 Jerusalem, Israel. Electronic address: shimon.schuldiner@huji.ac.il.
authors.forename: Shimon
authors.initials: S
authors.lastname: Schuldiner
mesh_terms: D000595:Amino Acid Sequence; D017920:Antiporters / Q000235:genetics / Q000378:metabolism*; D001407:Bacillus / Q000235:genetics; D001692:Biological Transport; D003001:Cloning, Molecular; D024901:Drug Resistance, Multiple, Bacterial; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000235:genetics; D010957:Plasmids; D012680:Sensitivity and Specificity; D016415:Sequence Alignment
chemical_list: D017920:Antiporters; D029968:Escherichia coli Proteins; C075436:EmrE protein, E coli
keywords: EmrE; antibiotic resistance; ion-coupled antiporters; methyl viologen; multidrug transporters