Liu J, Takiff H E, Nikaido H
Department of Molecular and Cell Biology, University of California, Berkeley, California 94720-3206, USA.
J Bacteriol. 1996 Jul;178(13):3791-5. doi: 10.1128/jb.178.13.3791-3795.1996.
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.
从耐喹诺酮耻垢分枝杆菌mc2-552染色体DNA中克隆的lfrA基因，当存在于多拷贝质粒上时，赋予对氟喹诺酮类药物低水平抗性。序列分析表明，lfrA编码主要转运体家族的膜外排泵（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）。在本研究中，我们研究了LfrA在耻垢分枝杆菌对氟喹诺酮类药物蓄积中的作用。携带lfrA基因质粒的耻垢分枝杆菌对亲水性喹诺酮诺氟沙星的稳态蓄积水平约为亲代菌株的50%，但通过添加质子导体羰基氰化物m-氯苯腙可将其提高至与亲代菌株相同的水平。LfrA介导的诺氟沙星外排受到环丙沙星的强烈竞争，但不受萘啶酸的竞争。此外，我们发现饥饿细胞蓄积的部分诺氟沙星在细胞重新获得能量后被泵出，并且这种外排速率在较高细胞内药物浓度下显示出饱和迹象。这些结果表明，LfrA多肽催化几种喹诺酮类药物的主动外排。
