title: Characterization of MATE-type multidrug efflux pumps from Klebsiella pneumoniae MGH78578.
abstract: We previously described the cloning of genes related to drug resistance from Klebsiella pneumoniae MGH78578. Of these, we identified a putative gene encoding a MATE-type multidrug efflux pump, and named it ketM. Escherichia coli KAM32 possessing ketM on a plasmid showed increased minimum inhibitory concentrations for norfloxacin, ciprofloxacin, cefotaxime, acriflavine, Hoechst 33342, and 4',6-diamidino-2-phenyl indole (DAPI). The active efflux of DAPI was observed in E. coli KAM32 possessing ketM on a plasmid. The expression of mRNA for ketM was observed in K. pneumoniae cells, and we subsequently disrupted ketM in K. pneumoniae ATCC10031. However, no significant changes were observed in drug resistance levels between the parental strain ATCC10031 and ketM disruptant, SKYM. Therefore, we concluded that KetM was a multidrug efflux pump, that did not significantly contribute to intrinsic resistance to antimicrobial chemicals in K. pneumoniae. MATE-type transporters are considered to be secondary transporters; therefore, we investigated the coupling cations of KetM. DAPI efflux by KetM was observed when lactate was added to produce a proton motive force, indicating that KetM effluxed substrates using a proton motive force. However, the weak efflux of DAPI by KetM was also noted when NaCl was added to the assay mixture without lactate. This result suggests that KetM may utilize proton and sodium motive forces.
authors: Department of Microbiology and Biochemistry, Daiichi University of Pharmacy, Fukuoka, Japan; Department of Microbiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University, Okayama, Japan.
authors: Wakano
authors: W
authors: Ogawa
authors: Department of Microbiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University, Okayama, Japan.
authors: Yusuke
authors: Y
authors: Minato
authors: Department of Microbiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University, Okayama, Japan.
authors: Hayata
authors: H
authors: Dodan
authors: Department of Microbiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University, Okayama, Japan.
authors: Motoyasu
authors: M
authors: Onishi
authors: Department of Microbiology, College of Pharmaceutical Sciences, Ritsumeikan University, Shiga, Japan.
authors: Tomofusa
authors: T
authors: Tsuchiya
authors: Department of Microbiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University, Okayama, Japan.
authors: Teruo
authors: T
authors: Kuroda
mesh_terms: D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics*; D003001:Cloning, Molecular / Q000379:methods; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics; D007711:Klebsiella pneumoniae / Q000235:genetics*; D026901:Membrane Transport Proteins / Q000235:genetics*; D008826:Microbial Sensitivity Tests / Q000379:methods; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D018892:Proton-Motive Force / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D016415:Sequence Alignment
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D026901:Membrane Transport Proteins; D012333:RNA, Messenger