Characterization of SFO-1, a Plasmid-Mediated Inducible Class A β-Lactamase from Enterobacter cloacae

Enterobacter cloacae 8009 produced an inducible class A β-lactamase which hydrolyzed cefotaxime efficiently. It also hydrolyzed other β-lactams except cephamycins and carbapenems. The activity was inhibited by clavulanic acid and imipenem. Thebla gene was transferable toEscherichia coli by electroporation of plasmid DNA. The molecular mass of the β-lactamase was 29 kDa and its pI was 7.3. All of these phenotypic characteristics of the enzyme except for inducible production resemble those of some extended-spectrum class A β-lactamases like FEC-1. The gene encoding this β-lactamase was cloned and sequenced. The deduced amino acid sequence of the β-lactamase was homologous to the AmpA sequences of theSerratia fonticola chromosomal enzyme (96%), MEN-1 (78%),Klebsiella oxytoca chromosomal enzymes (77%), TOHO-1 (75%), and FEC-1 (72%). The conserved sequences of class A β-lactamases, including the S-X(T)-X(S)-K motif, in the active site were all conserved in this enzyme. On the basis of the high degree of homology to the β-lactamase ofS. fonticola , the enzyme was named SFO-1. TheampR gene was located upstream of theampA gene, and the AmpR sequence of SFO-1 had homology with the AmpR sequences of the chromosomal β-lactamases fromCitrobacter diversus (80%),Proteus vulgaris (68%), andPseudomonas aeruginosa (60%). SFO-1 was also inducible inE. coli . However, a transformant harboring plasmid without intactampR produced a small amount of β-lactamase constitutively, suggesting that AmpR works as an activator ofampA of SFO-1. This is the first report from Japan describing an inducible plasmid-mediated class A β-lactamase in gram-negative bacteria.