Active efflux of fluoroquinolones in Mycobacterium smegmatis mediated by LfrA, a multidrug efflux pump.
The lfrA gene cloned from chromosomal DNA of quinolone-resistant Mycobacterium smegmatis mc2-552 conferred low-level resistance to fluoroquinolones when present on multicopy plasmids. Sequence analysis suggested that lfrA encodes a membrane efflux pump of the major facilitator family (H. E. Takiff, M. Cimino, M. C. Musso, T. Weisbrod, R. Martinez, M. B. Delgado, L Salazar, B. R. Bloom, and W. R. Jacbos, Jr., Proc. Natl. Acad. Sci. USA 93:362-366, 1996). In this work, we studied the role of LfrA in the accumulation of fluoroquinolones by M. smegmatis. The steady-state accumulation level of a hydrophilic quinolone, norfloxacin, by M. smegmatis harboring a plasmid carrying the lfrA gene was about 50% of that by the parent strain but was increased to the same level as that of the parent strain by addition of a proton conductor, carbonyl cyanide m-chorophenylhydrazone. Norfloxacin efflux mediated by LfrA was competed for strongly by ciprofloxacin but not by nalidixic acid. Furthermore, we showed that portions of norfloxacin accumulated by starved cells were pumped out upon reenergization of the cells, and the rates of this efflux showed evidence of saturation at higher intracellular concentrations of the drug. These results suggest that the LfrA polypeptide catalyzes the active efflux of several quinolones.
Liu
J
J
Department of Molecular and Cell Biology, University of California, Berkeley, California 94720-3206, USA.
Takiff
H E
HE
Nikaido
H
H
D000890:Anti-Infective Agents / Q000378:metabolism*; D017920:Antiporters / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins; D001693:Biological Transport, Active; D004352:Drug Resistance, Microbial; D018432:Drug Resistance, Multiple; D009161:Mycobacterium / Q000378:metabolism*; D009643:Norfloxacin / Q000378:metabolism*; D013379:Substrate Specificity
D000890:Anti-Infective Agents; D017920:Antiporters; D001426:Bacterial Proteins; C097688:LfrA protein, Mycobacterium smegmatis; D009643:Norfloxacin