ABSTRACT
The low susceptibility of enterococci to beta-lactams is due to the activity of the low-affinity penicillin-binding protein 5 (PBP5). One important feature of PBP5 is its ability to substitute for most, if not all, penicillin-binding proteins when they are inhibited. That substitution activity was analyzed in Enterococcus hirae SL2, a mutant whose pbp5 gene was interrupted by the nisRK genes and whose PBP3 synthesis was submitted to nisin induction. Noninduced SL2 cells were unable to divide except when plasmid-borne pbp5 genes were present, provided that the PBP5 active site was functional. Potential protein-protein interaction sites of the PBP5 N-terminal module were mutagenized by site-directed mutagenesis. The T167-L184 region (designated site D) appeared to be an essential intramolecular site needed for the stability of the protein. Mutations made in the two globular domains present in the N-terminal module indicated that they were needed for the suppletive activity. The P197-N209 segment (site E) in one of these domains seemed to be particularly important, as single and double mutations reduced or almost completely abolished, respectively, the action of PBP5.
