Inhibitor resistant class A beta-lactamases.
ABSTRACT Beta-lactamase inhibitors (clavulanic acid, tazobactam, and sulbactam) greatly enhance the therapeutic efficacy of their partner antibiotics (amoxacillin, ampicillin, piperacillin, and ticarcillin) against common enteric and non-enteric organisms possessing class A beta-lactamases. Unfortunately, the number of class A enzymes being discovered that are resistant to these combinations is increasingly rapidly. The TEM and SHV class A beta-lactamases resistant to inhibitors have point mutations in critical amino acids important for catalysis. Compared to the wild type beta-lactamase, inhibitor resistant enzymes are inefficient at hydrolyzing benzylpenicillin, aminopenicillins, and cephalosporins. Nevertheless, hyper-production of these enzymes resulting from mutations in the promoter region can confer substantial levels of resistance. Understanding the microbiologic and kinetic properties of these inhibitor resistant class A beta-lactamases can lead to the design of more potent beta-lactam compounds as well as more effective inhibitors.
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ABSTRACT: beta-Lactamases are extracellular or periplasmic bacterial enzymes which confer resistance to beta-lactam antibiotics. On the basis of their catalytic mechanisms, they can be divided into two major groups: active-site serine enzymes (classes A, C and D) and the ZnII enzymes (class B). The first crystal structure of a class B enzyme, the metallo-beta-lactamase from Bacillus cereus, has been solved at 2.5 A resolution [Carfi, Pares, Duée, Galleni, Duez, Frère & Dideberg (1995). EMBO J. 14, 4914-4921]. Recently, the crystal structure of the metallo-beta-lactamase from Bacteroides fragilis has been determined in a tetragonal space group [Concha, Rasmussen, Bush & Herzberg (1996). Structure, 4, 823-836]. The structure of the metallo-beta-lactamase from B. fragilis in an orthorhombic crystal form at 2.0 A resolution is reported here. The final crystallographic R is 0.196 for all the 32501 observed reflections in the range 10-2.0 A. The refined model includes 458 residues, 437 water molecules, four zinc and two sodium ions. These structures are discussed with reference to Zn binding and activity. A catalytic mechanism is proposed which is coherent with metallo-beta-lactamases being active with either one Zn ion (as in Aeromonas hydrophila) or two Zn ions (as in B. fragilis) bound to the protein.Acta Crystallographica Section D Biological Crystallography 01/1998; 54(1):45-57. · 14.10 Impact Factor
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ABSTRACT: DNA-DNA hybridization and sequencing were performed to determine the molecular basis of resistance to clavulanic acid in 107 inhibitor-resistant TEM (IRT) enzymes produced by Escherichia coli clinical isolates. These beta-lactamases derived from TEM-1 enzyme focused at pI 5.2 (n = 68) or 5.4 (n = 39) and were very poorly inhibited by clavulanic acid compared with TEM-1 enzyme. Results showed that the amino acid sequences of 84 of the 107 enzymes differ from TEM-1 by one or two substitutions previously described: Arg-244-->Ser (IRT-2) in 22 strains, Met-69-->Leu (TEM-33) in 17 strains, Met-69-->Val (TEM-34) in 14 strains, Met-69-->Ile (IRT-3) in 6 strains, Met-69-->Leu associated with Asn-276-->Asp (IRT-4) in 13 strains, and Met-69-->Val associated with Asn-276-->Asp (TEM-36) in 12 strains. A new combination, Met-69-->Ile with Asn-276-->Asp, was found in 20 strains and was called IRT-8. Two IRT enzymes not previously described were characterized. The substitution Met-69-->Val associated with a novel substitution Arg-275-->Leu occurred in one strain. The combination Met-69-->Leu and Asn-276-->Asp was associated with the novel substitution Trp-165-->Arg in two strains. These two novel enzymes were called IRT-9 and IRT-10, respectively. The implication of these novel mutated positions, 165 and 275, in resistance to inactivation by clavulanate was supported by crystallographic data on the TEM-1 enzyme and results of site-directed mutagenesis. Molecular characterization of these mutants showed great diversity among the genes coding for inhibitor-resistant TEM enzymes produced by clinical E. coli isolates.Antimicrobial Agents and Chemotherapy 03/1995; 39(2):427-30. · 4.57 Impact Factor
- Antimicrobial Agents and Chemotherapy 07/1995; 39(6):1211-33. · 4.57 Impact Factor