OHIO-1 beta-lactamase mutants: Asp179Gly mutation confers resistance to ceftazidime

FEMS Microbiol Lett. 1997 Jul 15;152(2):275-8. doi: 10.1111/j.1574-6968.1997.tb10439.x.

Abstract

The Asp179Gly mutant of the OHIO-1 beta-lactamase, an SHV enzyme, was constructed to investigate the effect of disruption of the omega loop on beta-lactamase activity in this class A enzyme. In Escherichia coli DH5 alpha the strain possessing the Asp179Gly mutation of the OHIO-1 beta-lactamase demonstrated increased susceptibility to all beta-lactams except ceftazidime and ceftriaxone. The minimum inhibitory concentrations for ceftazidime and ceftriaxone increased from 0.25 and 0.015 microgram/ml to 4.0 and .25 micrograms/ ml respectively. For ceftazidime, a substrate not hydrolyzed by the wild-type enzyme (K(m) > or = 500 microM), the K(m) of the Asp179Gly mutant beta-lactamase was measured to be 7 microM and the Vmax was 0.13 microM/min. The minimum inhibitory concentrations, K(m), and Vmax for all other beta-lactams decreased. Our analysis of this OHIO-1 beta-lactamase mutant suggests that disruption of the salt bridge in the omega loop by substitution of a glycine at position 179 markedly decreases the catalytic efficiency of the enzyme.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Aspartic Acid / physiology*
  • Ceftazidime / pharmacology*
  • Cephalosporin Resistance / genetics*
  • Cephalosporins / pharmacology*
  • Escherichia coli / drug effects
  • Kinetics
  • Microbial Sensitivity Tests
  • Mutation
  • Penicillins / pharmacology
  • beta-Lactamases / chemistry
  • beta-Lactamases / genetics*

Substances

  • Cephalosporins
  • Penicillins
  • Aspartic Acid
  • Ceftazidime
  • beta-Lactamases