A PHOSPHOGLUCOMUTASE-LIKE GENE ESSENTIAL FOR THE OPTIMAL EXPRESSION OF METHICILLIN RESISTANCE IN STAPHYLOCOCCUS-AUREUS - MOLECULAR-CLONING AND DNA-SEQUENCING

We describe here the cloning and sequencing of a new auxiliary gene id entified by Tn551 insertional mutagenesis of the highly and homogeneou sly methicillin-resistant Staphylococcus aureus strain COL, The insert ionally inactivated mutant RUSA315 had intact mecA and normal amounts of PBP2A, but drastically reduced antibiotic resistance (drop in methi cillin MIC from 1600 to 1.5 mu g ml(-1)), a unique heterogeneous pheno type, and a compositional change in the cell wall characterized by the complete disappearance of the unsubstituted disaccharide pentapeptide from the peptidoglycan, Cloning in E, coil followed by sequencing loc ated the Tn551 insert Omega 720 in an open reading frame of 451 codons , provisionally called femR315, defining a polypeptide with a deduced amino acid sequence that showed over 26% sequence identity and 57% ove rall sequence similarity with the phosphoglucomutase (PGM) gene of E, coli, The Tn551 insertion site of a previously described mutant 12F (f emD) also lies in the same gene as femR315, The wild-type form of femR 315 subcloned in a shuttle vector fully restored expression of high le vel (parental) methicillin resistance in mutant RUSA315, The exact bio chemical function of femR315 is not known, However, enzymes similar to PGM catalyze the isomerization of hexose and hexosamine phosphates le ading to the formation of glucosamine-1-P, which is an obligate precur sor in the biosynthesis of UDP-N-acetylglucosamine (UDP-NAGA). We prop ose that the suppression of methicillin resistance in RUSA315 is relat ed to some functional or quantitative abnormality of UDP-NAGA metaboli sm.