Chromosomal β-lactamase of Klebsiella oxytoca, a new class A enzyme that hydrolyzes broad-spectrum β-lactam antibiotics

The chromosomally encoded β-lactamase gene of Klebsiella oxytoca E23004, a strain resistant to cefoperazone and aztreonam, was cloned and expressed in Escherichia coli HB101. The molecular mass and pI of this enzyme were 28 kilodaltons and 7.4, respectively. Although the β-lactamase of K. oxytoca hydrolyzed many cephalosporins, including broad-spectrum drugs, the nucleotide sequence and deduced amino acid sequence lacked homology with chromosomal class C β-lactamase genes (ampC) of E. coli or Citrobacter freundii. Rather, about 45% nucleotide sequence homology and 40% deduced amino acid sequence homology were observed between the K. oxytoca β-lactamase and TEM-1, a class A β-lactamase which does not efficiently hydrolyze cephalosporins. Values of K(m), relative V(max), and relative V(max)/K(m) for the K. oxytoca β-lactamase indicated that the enzyme is a penicillinase but that it can hydrolyze cefoperazone effectively and other broad-spectrum cephems weakly. Hence, the chromosomal β-lactamase of K. oxytoca E23004 belongs to class A but differences in its amino acid sequence provide a broader spectrum of activity.