1. Biochemical characterization of the FEZ-1 metallo-beta-lactamase of Legionella gormanii ATCC 33297T produced in Escherichia coli.
- Author
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Mercuri PS, Bouillenne F, Boschi L, Lamotte-Brasseur J, Amicosante G, Devreese B, van Beeumen J, Frère JM, Rossolini GM, and Galleni M
- Subjects
- Amino Acid Sequence, Binding Sites, Cephalosporin Resistance, Chelating Agents pharmacology, Escherichia coli genetics, Genes, Bacterial, Hydrogen-Ion Concentration, Kinetics, Models, Molecular, Molecular Sequence Data, Sequence Homology, Amino Acid, Transfection, Zinc analysis, beta-Lactamases metabolism, Legionella enzymology, Legionella genetics, beta-Lactamases chemistry, beta-Lactamases genetics
- Abstract
The bla(FEZ-1) gene coding for the metallo-beta-lactamase of Legionella (Fluoribacter) gormanii ATCC 33297T was overexpressed via a T7 expression system in Escherichia coli BL21(DE3)(pLysS). The product was purified to homogeneity in two steps with a yield of 53%. The FEZ-1 metallo-beta-lactamase exhibited a broad-spectrum activity profile, with a preference for cephalosporins such as cephalothin, cefuroxime, and cefotaxime. Monobactams were not hydrolyzed. The beta-lactamase was inhibited by metal chelators. FEZ-1 is a monomeric enzyme with a molecular mass of 29,440 Da which possesses two zinc-binding sites. Its zinc content did not vary in the pH range of 5 to 9, but the presence of zinc ions modified the catalytic efficiency of the enzyme. A model of the FEZ-1 three-dimensional structure was built.
- Published
- 2001
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