Comparative analysis of Pseudomonas aeruginosa penicillin-binding protein 7 in the context of its membership in the family of low-molecular-mass PBPs.

The Pseudomonas aeruginosa pbpG gene encoding penicillin-binding protein 7, a homologue of the Escherichia coli gene encoding a DD-endopeptidase, was cloned and sequenced, pbpG was located immediately downstream of the phenylalanine hydroxylase (phh) operon. DNA sequencing revealed an open reading frame of 936 bp (starting with a GTG codon) which encodes a protein of 34,115 Da. N-terminal amino acid sequencing confirmed the presence of a cleavable N-terminal signal peptide of 23 amino acids. Verification that the protein is a penicillin-binding protein was directly demonstrated by labelling with 125I-labelled penicillin X. Inactivation of P. aeruginosa pbpG by interposon mutagenesis resulted in no obvious phenotypic changes, but when P. aeruginosa PbpG was overexpressed in E. coli using a T7 expression system, cell lysis resulted. P. aeruginosa PbpG resembled E. coli PbpG in being associated with the membrane fraction. Two additional members of the PbpG subfamily were identified in the database. P. aeruginosa PbpG shows 63% identity with E. coli penicillin-binding protein 7 (PbpG) and 60% identity with Vibrio cholerae PbpG, but only 23% identity with Haemophilus influenzae PbpG. The PbpG subfamily and three other subfamilies constituting the low-molecular-mass PBP protein family were analysed by multiple alignment of 26 sequences. PbpG exhibited the consensus motifs of other penicillin-binding proteins. Ten anchor residues were identified that are conserved at the family level within the superfamily of serine-active-site penicillin-interacting proteins.