Sequence analysis of PER-1 extended-spectrum beta-lactamase from Pseudomonas aeruginosa and comparison with class A beta-lactamases

Abstract

We have determined the nucleotide sequence (EMBL accession number, Z 21957) of the cloned chromosomal PER-1 extended-spectrum beta-lactamase gene from a Pseudomonas aeruginosa RNL-1 clinical isolate, blaPER-1 corresponds to a 924-bp open reading frame which encodes a polypeptide of 308 amino acids. This open reading frame is preceded by a -10 and a -35 region consistent with a putative P. aeruginosa promoter. Primer extension analysis of the PER-1 mRNA start revealed that this promoter was active in P. aeruginosa but not in Escherichia coli, in which PER-1 expression was driven by vector promoter sequences. N-terminal sequencing identified the PER-1 26-amino-acid leader peptide and enabled us to calculate the molecular mass (30.8 kDa) of the PER-1 mature form. Analysis of the percent GC content of blaPER-1 and of its 5' upstream sequences, as well as the codon usage for blaPER-1, indicated that blaPER-1 may have been inserted into P. aeruginosa genomic DNA from a nonpseudomonad bacterium. The PER-1 gene showed very low homology with other beta-lactamase genes at the DNA level. By using computer methods, assessment of the extent of identity between PER-1 and 10 beta-lactamase amino acid sequences indicated that PER-1 is a class A beta-lactamase. PER-1 shares around 27% amino acid identity with the sequenced extended-spectrum beta-lactamases of the TEM-SHV series and MEN-1 from Enterobacteriaceae species. The use of parsimony methods showed that PER-1 is not more closely related to gram-negative than to gram-positive bacterial class A beta-lactamases. Surprisingly, among class A beta-lactamases, PER-1 was most closely related to the recently reported CFXA from Bacteroides vulgatus, with which it shared 40% amino acid identity. This work indicates that non-Enterobacteriaceae species such as P. aeruginosa may possess class A extended-spectrum beta-lactamase genes possibly resulting from intergeneric DNA transfer.