A synergistic effect between vancomycin or teicoplanin and different beta-lactam antibiotics was found for two strains of Enterococcus faecium, EFM4 and EFM11, expressing resistance to glycopeptides and belonging to the VANA class. The MICs of penicillin for these two strains were 16 and 128 micrograms/ml, respectively. By using a penicillin-binding protein (PBP) competition assay, it was shown that the affinities of PBPs for different beta-lactam antibiotics and the MICs of these antibiotics obtained in the presence of teicoplanin correlated with the substitution of two high-molecular-weight PBPs for the low-molecular-weight PBP5 as the essential target. Mutants of EFM4 and EFM11 which had lost the synergistic effect between beta-lactams and glycopeptides were selected on teicoplanin plus ceftriaxone at a frequency of 10(-5) and 10(-3), respectively. The mechanism of the loss of synergy was explored. For the mutants derived from EFM4, it was associated with a change in PBPs, while for the mutants derived from EFM11, it was related to some unknown change on the conjugative plasmid responsible for the glycopeptide resistance. These combined observations reflect the relationship which seems to exist between the new D-lactate peptidoglycan precursor, synthesized when the vancomycin resistance is expressed, and the affinity of the different PBPs for this precursor.