title: Functional effects of intramembranous proline substitutions in the staphylococcal multidrug transporter QacA.
abstract: The QacA multidrug transporter is encoded on Staphylococcus aureus multidrug resistance plasmids and confers broad-range antimicrobial resistance to more than 30 monovalent and bivalent lipophilic, cationic compounds from at least 12 different chemical classes. QacA contains 10 proline residues predicted to be within transmembrane regions, several of which are conserved in related export proteins. Proline residues are classically known as helix-breakers and are highly represented within the transmembrane helices of membrane transport proteins, where they can mediate the formation of structures essential for protein stability and transport function. The importance of these 10 intramembranous proline residues for QacA-mediated transport function was determined by examining the functional effect of substituting these residues with glycine, alanine or serine. Several proline-substituted QacA mutants failed to confer high-level resistance to selected QacA substrates. However, no single proline mutation, including those at conserved positions, significantly disrupted QacA protein expression or QacA-mediated resistance to all representative substrates, suggesting that these residues are not essential for the formation of structures requisite to the QacA substrate transport mechanism.
authors.affiliation: School of Biological Sciences, University of Sydney, Sydney, NSW, Australia.
authors.forename: Karl A
authors.initials: KA
authors.lastname: Hassan
authors.forename: Melanie
authors.initials: M
authors.lastname: Galea
authors.forename: Jingqin
authors.initials: J
authors.lastname: Wu
authors.forename: Bernadette A
authors.initials: BA
authors.lastname: Mitchell
authors.forename: Ronald A
authors.initials: RA
authors.lastname: Skurray
authors.forename: Melissa H
authors.initials: MH
authors.lastname: Brown
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000900:Anti-Bacterial Agents / Q000378:metabolism / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015153:Blotting, Western; D024901:Drug Resistance, Multiple, Bacterial; D026901:Membrane Transport Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011392:Proline / Q000378:metabolism*; D016415:Sequence Alignment; D013211:Staphylococcus aureus / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D026901:Membrane Transport Proteins; C068498:qacA protein, Staphylococcus aureus; D011392:Proline