SITE-DIRECTED MUTAGENESIS OF THE MECA GENE FROM A METHICILLIN-RESISTANT STRAIN OF STAPHYLOCOCCUS-AUREUS

The mecA-27r gene from Staphylococcus aureus 27r encodes penicillin-bi nding protein 2a (PBP2a-27r), which causes this strain to be methicill in resistant. Removal or replacement of the N-terminal transmembrane d omain had no effect on binding of penicillin, but removal of portions of the putative transglycosylase domain (144, 245, or 341 amino acids after the transmembrane region) destroyed penicillin-binding activity. The SXXK, SXN, and KSG motifs, present in all penicillin-interacting enzymes, were found in the expected linear spatial arrangement within the putative transpeptidase region of PBP2a-27r. Alterations of amino acids in all three of these motifs resulted in elimination of penicill in-binding activity, confirming their roles in the interaction with pe nicillin.