title: PatA and PatB form a functional heterodimeric ABC multidrug efflux transporter responsible for the resistance of Streptococcus pneumoniae to fluoroquinolones.
abstract: All bacterial multidrug ABC transporters have been shown to work as either homodimers or heterodimers. Two possibly linked genes, patA and patB from Streptococcus pneumococcus, that encode half-ABC transporters have been shown previously to be involved in fluoroquinolone resistance. We showed that the ΔpatA, ΔpatB, or ΔpatA/ΔpatB mutant strains were more sensitive to unstructurally related compounds, i.e., ethidium bromide or fluoroquinolones, than the wild-type R6 strain. Inside-out vesicles prepared from Escherichia coli expressing PatA and/or PatB transported Hoechst 33342, a classical substrate of multidrug transporters, only when both PatA and PatB were coexpressed. This transport was inhibited either by orthovanadate or by reserpine, and mutation of the conserved Walker A lysine residue of either PatA or PatB fully abrogated Hoechst 33342 transport. PatA, PatB, and the PatA/PatB heterodimer were purified from detergent-solubilized E. coli membrane preparations. Protein dimers were identified in all cases, albeit in different proportions. In contrast to the PatA/PatB heterodimers, homodimers of PatA or PatB failed to show a vanadate-sensitive ATPase activity. Thus, PatA and PatB need to interact together to make a functional drug efflux transporter, and they work only as heterodimers.
authors.affiliation: Université Joseph Fourier-Grenoble 1, Institut de Biologie Structurale, Grenoble, France.
authors.forename: Emilie
authors.initials: E
authors.lastname: Boncoeur
authors.forename: Claire
authors.initials: C
authors.lastname: Durmort
authors.forename: Benoît
authors.initials: B
authors.lastname: Bernay
authors.forename: Christine
authors.initials: C
authors.lastname: Ebel
authors.forename: Anne Marie
authors.initials: AM
authors.lastname: Di Guilmi
authors.forename: Jacques
authors.initials: J
authors.lastname: Croizé
authors.forename: Thierry
authors.initials: T
authors.lastname: Vernet
authors.forename: Jean-Michel
authors.initials: JM
authors.lastname: Jault
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000900:Anti-Bacterial Agents / Q000378:metabolism / Q000494:pharmacology*; D024881:Drug Resistance, Bacterial; D024841:Fluoroquinolones / Q000378:metabolism / Q000494:pharmacology*; D017353:Gene Deletion; D006801:Humans; D009154:Mutation; D011008:Pneumococcal Infections / Q000188:drug therapy; D055503:Protein Multimerization; D013296:Streptococcus pneumoniae / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D014638:Vanadates / Q000378:metabolism
chemical_list: D018528:ATP-Binding Cassette Transporters; D000900:Anti-Bacterial Agents; D024841:Fluoroquinolones; D014638:Vanadates