Molecular cloning and biochemical characterization of VIM-12, a novel hybrid VIM-1/VIM-2 metallo-beta-lactamase from a Klebsiella pneumoniae clinical isolate, reveal atypical substrate specificity.
Metallo-beta-lactamases (MBLs) are considered an emerging family of Zn2+-dependent enzymes that significantly contribute to the resistance of many nosocomial pathogens against beta-lactam antimicrobials. Since these plasmid-encoded enzymes constitute specific molecular targets for beta-lactams, their exact mode of action is greatly important in deploying efficient anti-infective treatments and for the control of severe multi-resistant nosocomial infections, which becomes a global problem. A novel hybrid VIM-1/VIM-2-type beta-lactamase (named VIM-12) has recently been identified in a clinical isolate of Klebsiella pneumoniae in Greece. The sequence of this enzyme is highly similar with that of VIM-1 at its N-terminal region and with that of VIM-2 at its C-terminal region, raising the question of whether this sequence similarity reflects also a similar functional role. Moreover, the possible contribution of this novel beta-lactamase to the overall antibiotic resistance of this specific clinical isolate was investigated. The gene encoding VIM-12 was cloned and expressed, and the recombinant enzyme was used for detailed kinetic analysis, using a variety of beta-lactam antibiotics. VIM-12 was found to exhibit narrow substrate specificity, compared to other known beta-lactamases, limited mainly to penicillin and to a much lesser extent to imipenen. Interestingly, meropenem was found to act as a noncompetitive inhibitor of the enzyme, although the active site of VIM-12 exhibited complete conservation of residues among VIM enzymes. We conclude that VIM-12 represents a novel and unique member of the family of known metallo-beta-lactamases, exhibiting atypical substrate specificity.
Kontou
Maria
M
Department of Biochemistry and Biotechnology, University of Thessaly, 26 Ploutonos st., 41221 Larissa, Greece.
Pournaras
Spyros
S
Kristo
Ioulia
I
Ikonomidis
Alexandros
A
Maniatis
Antonios N
AN
Stathopoulos
Constantinos
C
D000368:Aged; D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D007710:Klebsiella Infections / Q000382:microbiology; D007711:Klebsiella pneumoniae / Q000201:enzymology*; D008969:Molecular Sequence Data; D016415:Sequence Alignment; D013379:Substrate Specificity; D001618:beta-Lactamases / Q000235:genetics / Q000378:metabolism*
D001618:beta-Lactamases; C525149:beta-lactamase VIM-12, Klebsiella pneumoniae