Functional cloning and expression of emeA, and characterization of EmeA, a multidrug efflux pump from Enterococcus faecalis.
A fragment of chromosomal DNA from Enterococcus faecalis ATCC 29212 was cloned using Escherichia coli KAM32 host cells lacking major multidrug efflux pumps. E. coli KAM32 cells were sensitive to many antimicrobial agents, and the transformed cells harboring a recombinant plasmid became resistant to several structurally unrelated antimicrobial agents such as tetraphenylphosphonium chloride, 4',6-diamidino-2-phenylindole (DAPI), Hoechst 33342, acriflavine, benzalkonium chloride, norfloxacin and ethidium bromide. This suggests that the cloned DNA fragment carries a gene(s) encoding a multidrug efflux pump. Determination of the nucleotide sequence of the cloned DNA revealed a gene designated as emeA. The transformed E. coli cells showed efflux activity of several antimicrobial agents such as DAPI, Hoechst 33342 and acriflavine. Efflux of DAPI via EmeA was strongly inhibited by reserpine.
Lee
Eun-Woo
EW
Department of Microbiology, Faculty of Pharmaceutical Sciences, Okayama University, Okayama , Japan.
Chen
Jing
J
Huda
Md Nazmul
MN
Kuroda
Teruo
T
Mizushima
Tohru
T
Tsuchiya
Tomofusa
T
D001426:Bacterial Proteins; D003001:Cloning, Molecular / Q000379:methods*; D013293:Enterococcus faecalis / Q000235:genetics* / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation / Q000502:physiology; D019450:Genes, MDR / Q000502:physiology; D026901:Membrane Transport Proteins / Q000096:biosynthesis* / Q000235:genetics*
D001426:Bacterial Proteins; C445688:EmeA protein, Enterococcus faecalis; D026901:Membrane Transport Proteins