Cloning and sequencing of the low-affinity penicillin-binding protein 3 super(r)-encoding gene of Enterococcus hirae) S185: Modular design and structural organization of the protein

The clinical isolate Enterococcus hirae S185 has a peculiar mode of resistance to penicillin in that it possesses two low-affinity penicillin-binding proteins (PBPs): the 71-kDa PBP5, also found in other enterococci, and the 77-kDa PBP3 super(r). The two PBPs have the same low affinity for the drug and are immunochemically related to each other. The PBP3 super(r)-encoding gene has been cloned and sequenced, and the derived amino acid sequence has been compared by computer-assisted hydrophobic cluster analysis with that of the low-affinity PBP5 of E. hirae) R40, the low-affinity PBP2' of Staphylococcus aureus , and the PBP2 of Escherichia coli used as the standard of reference of the high-M sub(r) PBPs of class B. On the basis of the shapes, sizes, and distributions of the hydrophobic and nonhydrophobic clusters along the sequences and the linear amino acid alignments derived from this analysis, the dyad PBP3 super(r)-PBP5 has an identity index of 78.5%, the triad PBP3 super(r)-PBP5-PBP2' has an identity index of 29%, and the tetrad PBP3 super(r)-PBP5-PBP2'-PBP2 (of E. coli)) has an identity index of 13%. In spite of this divergence, the low-affinity PBPs are of identical modular design and possess the nine amino acid groupings (boxes) typical of the N-terminal and C-terminal domains of the high-M sub(r) PBPs of class B. At variance with the latter PBPs, however, the low-affinity PBPs have an additional approximately equals 110-amino-acid polypeptide stretch that is inserted between the amino end of the N-terminal domain and the carboxy end of the membrane anchor. While the enterococcal PBP5 gene is chromosome borne, the PBP3 super(r) gene appears to be physically linked to the erm gene, which confers resistance to erythromycin and is known to be plasmid borne in almost all the Streptococcus) spp. examined.