CLONING OF A GENE FROM ESCHERICHIA-COLI THAT CONFERS RESISTANCE TO FOSMIDOMYCIN AS A CONSEQUENCE OF AMPLIFICATION

A gene conferring resistance to fosmidomycin (Fs) was cloned from the gene pool of a wild-type strain of Escherichia coil. The cloned DNA fr agment was sequenced and shown to encode a putative polypeptide of 406 amino acids (aa) with a molecular weight of 43 303. The gene mapped a t 10.9 min on the E. coli chromosome and was designated fsr (fosmidomy cin resistance). Maxicell analysis revealed that the Fsr protein migra ted in sodium dodecyl sulfate-polyacrylamide-gel electrophoresis as a broad band of 35 kDa. A comparison between the aa sequence of Fsr and sequences in a protein database revealed 18% homology to the bacterial drug-export proteins that mediate resistance to tetracycline and chlo ramphenicol. Hydropathy analysis of the Fsr protein revealed twelve pu tative transmembrane segments. The degree of Fs(R) of transformants de pended on the number of copies of the plasmid that contained fsr. The levels of ubiquinone-8 and undecaprenyl phosphate in cells that harbor ed a high-copy-number plasmid that included fsr were almost the same a s those in the cells without the plasmid. These results suggest that F sr does not have any direct effect on the biosynthesis of isoprenoid i n E. coli, and that the mechanism for Fs(R) involves the efflux of the drug by a process that is facilitated by Fsr.