BINDING AFFINITIES OF BETA-LACTAM ANTIBODIES FOR PENICILLIN-BINDING PROTEIN-2' IN METHICILLIN-RESISTANT STAPHYLOCOCCUS-AUREUS

We devised an accurate procedure with which to measure the affinities of beta-lactam antibiotics for penicillin-binding protein (PBP) 2' in methicillin-resistant Staphylococcus aureus (MRSA). In the present stu dy, we used two isogenic strains of MRSA, one heterogeneous and the ot her homogeneous, derived from the methicillin-susceptible strain FDA20 9P, harbouring the mecA gene. In these MRSA strains, PBP2' was saturat ed by [C-14]benzylpenicillin (PCG) at a concentration of 300 mg/L. In addition, the saturation of PBP2' by [C-14]PCG required an incubation period of 30 min. According to these results, the precise affinities o f beta-lactam antibiotics for PBP2' were determined by the 'accurate c ompetition assay', using a high concentration of [C-14]PCG and extendi ng the reaction time. This procedure yielded lower IC50 values of beta -lactams than the 'usual competition assay'. However, each beta-lactam had almost the same affinity for PBP2' in heterogeneous and homogeneo us strains. These results suggest there is a factor(s) other than PBP2 ' responsible for controlling resistance levels and the heterogeneity or homogeneity of MRSA strains.