The Structure of the Dizinc Subclass B2 Metallo-β-Lactamase CphA Reveals that the Second Inhibitory Zinc Ion Binds in the Histidine Site
- 1 October 2009
- journal article
- Published by American Society for Microbiology in Antimicrobial Agents and Chemotherapy
- Vol. 53 (10), 4464-4471
- https://doi.org/10.1128/aac.00288-09
Abstract
Bacteria can defend themselves against β-lactam antibiotics through the expression of class B β-lactamases, which cleave the β-lactam amide bond and render the molecule harmless. There are three subclasses of class B β-lactamases (B1, B2, and B3), all of which require Zn 2+ for activity and can bind either one or two zinc ions. Whereas the B1 and B3 metallo-β-lactamases are most active as dizinc enzymes, subclass B2 enzymes, such as Aeromonas hydrophila CphA, are inhibited by the binding of a second zinc ion. We crystallized A. hydrophila CphA in order to determine the binding site of the inhibitory zinc ion. X-ray data from zinc-saturated crystals allowed us to solve the crystal structures of the dizinc forms of the wild-type enzyme and N220G mutant. The first zinc ion binds in the cysteine site, as previously determined for the monozinc form of the enzyme. The second zinc ion occupies a slightly modified histidine site, where the conserved His118 and His196 residues act as metal ligands. This atypical coordination sphere probably explains the rather high dissociation constant for the second zinc ion compared to those observed with enzymes of subclasses B1 and B3. Inhibition by the second zinc ion results from immobilization of the catalytically important His118 and His196 residues, as well as the folding of the Gly232-Asn233 loop into a position that covers the active site.Keywords
This publication has 36 references indexed in Scilit:
- Trapping and Characterization of a Reaction Intermediate in Carbapenem Hydrolysis by B. cereus Metallo-β-lactamaseJournal of the American Chemical Society, 2008
- Engineered Mononuclear Variants in Bacillus cereus Metallo-β-lactamase BcII Are InactiveBiochemistry, 2008
- Competitive Inhibitors of the CphA Metallo-β-Lactamase from Aeromonas hydrophilaAntimicrobial Agents and Chemotherapy, 2007
- Mechanistic Studies on the Mononuclear ZnII-Containing Metallo-β-lactamase ImiS from Aeromonas sobriaBiochemistry, 2006
- Crystal Structure of Pseudomonas aeruginosa SPM-1 Provides Insights into Variable Zinc Affinity of Metallo-β-lactamasesJournal of Molecular Biology, 2006
- A Metallo-β-lactamase Enzyme in Action: Crystal Structures of the Monozinc Carbapenemase CphA and its Complex with BiapenemJournal of Molecular Biology, 2005
- Coot: model-building tools for molecular graphicsActa Crystallographica Section D-Biological Crystallography, 2004
- The crystal structure of the L1 metallo-β-lactamase from Stenotrophomonas maltophilia at 1.7 å resolutionJournal of Molecular Biology, 1998
- Refinement of Macromolecular Structures by the Maximum-Likelihood MethodActa Crystallographica Section D-Biological Crystallography, 1997
- The CCP4 suite: programs for protein crystallographyActa Crystallographica Section D-Biological Crystallography, 1994