title: Outer membrane protein OmpW participates with small multidrug resistance protein member EmrE in quaternary cationic compound efflux.
abstract: In Escherichia coli, the small multidrug resistance (SMR) transporter protein EmrE confers host resistance to a broad range of toxic quaternary cation compounds (QCC) via proton motive force in the plasma membrane. Biologically produced QCC also act as EmrE osmoprotectant substrates within the cell and participate in host pH regulation and osmotic tolerance. Although E. coli EmrE is one of the most well-characterized SMR members, it is unclear how the substrates it transports into the periplasm escape across the outer membrane (OM) in Gram-negative bacteria. We tested the hypothesis that E. coli EmrE relies on an unidentified OM protein (OMP) to complete the extracellular release of its QCC. Eleven OMP candidates were screened using an alkaline phenotypic growth assay to identify OMP involvement in EmrE-mediated QCC efflux. E. coli single-gene deletion strains were transformed with plasmid-carried copies of emrE to detect reduced-growth and rescued-growth phenotypes under alkaline conditions. Among the 11 candidates, only the ΔompW strain showed rescued alkaline growth tolerance when transformed with pEmrE, supporting the corresponding protein's involvement in EmrE osmoprotectant efflux. Coexpression of plasmids carrying the ompW and emrE genes transformed into the E. coli ΔompW and ΔemrE strains demonstrated a functional complementation restoring the original alkaline loss-of-growth phenotype. Methyl viologen drug resistance assays of pEmrE and pOmpW plasmid-complemented E. coli ΔompW and wild-type strains found higher host drug resistance than with other plasmid combinations. This study confirms our hypothesis that the porin OmpW participates in the efflux of EmrE-specific substrates across the OM.
authors.affiliation: Department of Biological Sciences, University of Calgary, Calgary, Alberta, Canada.
authors.forename: Maria S
authors.initials: MS
authors.lastname: Beketskaia
authors.forename: Denice C
authors.initials: DC
authors.lastname: Bay
authors.forename: Raymond J
authors.initials: RJ
authors.lastname: Turner
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D017920:Antiporters / Q000235:genetics / Q000378:metabolism*; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D024901:Drug Resistance, Multiple, Bacterial; D004926:Escherichia coli / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D008826:Microbial Sensitivity Tests; D013379:Substrate Specificity
chemical_list: D000900:Anti-Bacterial Agents; D017920:Antiporters; D001425:Bacterial Outer Membrane Proteins; D029968:Escherichia coli Proteins; C064035:ompW protein, E coli; C075436:EmrE protein, E coli