MOLECULAR AND GENETIC-ANALYSIS OF THE BACTEROIDES UNIFORMIS CEPHALOSPORINASE GENE, CBLA, ENCODING THE SPECIES-SPECIFIC BETA-LACTAMASE

The gene, cblA, encoding the species-specific, clavulanate-susceptible , endogenous cephalosporinase was cloned from Bacteroides uniformis WA L-7088. The nucleotide sequence was determined, and the cblA structura l gene was found to be 891 nucleotides, with a 48% G+C composition, wh ich is similar to that of the B. uniformis genome. The cblA open readi ng frame encoded an Ambler class A beta-lactamase polypeptide precurso r of 296 amino acid residues with a predicted molecular weight of 33,4 50. A beta-lactamase-deficient B. uniformis mutant with increased beta -lactam susceptibility was constructed by insertional inactivation of the chromosomal gene. This mutant was complemented by plasmids bearing the cblA gene, and the resulting strains were resistant to cephalorid ine and had a beta-lactamase that comigrated with the parental beta-la ctamase on sodium dodecyl sulfate-polyacrylamide gel electrophoresis ( 30,500 Da) and in isoelectric focusing gels (pI 4.6), confirming a rol e for this beta-lactamase in resistance.