Characterization and amino acid sequence of the OXY-2 group β-lactamase of pI 5.7 isolated from aztreonam-resistant Klebsiella oxytoca strain HB60

Abstract

Klebsiella oxytoca strain HB60 is highly resistant to cefoperazone and aztreonam (MICs = 128 mg/L). It produces a chromosomally encoded β-lactamase of pI 5.7 which was highly efficient against penicillins, first-generation cephalosporins and cefoperazone, a non-oxyimino third-generation cephalosporin. Aztreonam and oxyimino broad-spectrum cephalosporins were less good substrates. The β-lactamase activity was susceptible to inhibition by clavulanic acid (IC50 = 1 μM). The enzyme purified to homogeneity had a specific activity towards benzylpenicillin of 3670 U/mg. The 263 amino acid residues of the protein were sequenced by Edman degradation of proteolytic peptides. The β-lactamase was shown to belong to the OXY-2 group as it had only one amino acid substitution (Asn for Asp at ABL position 197) compared with the β-lactamase (pI 5.2) from the aztreonam-susceptible K. oxytoca strain SL911 and two substitutions (Ala223 for Val and Asp255 for Asn) compared with the β-lactamase (pI 6.4) from the aztreonam-resistant K. oxytoca strain D488. These three OXY-2-group enzymes behave in the same way towards β-lactam antibiotics. The variability in the resistance of these K. oxytoca strains would thus seem to be due to variation in the level of production of the β-lactamases rather than to structural alteration of the enzymes.