Structure of GES-1 at atomic resolution: insights into the evolution of carbapenamase activity in the class A extended-spectrum beta-lactamases

Acta Crystallogr D Biol Crystallogr. 2007 Sep;63(Pt 9):982-92. doi: 10.1107/S0907444907036955. Epub 2007 Aug 17.

Abstract

The structure of the class A extended-spectrum beta-lactamase GES-1 from Klebsiella pneumoniae has been determined to 1.1 A resolution. GES-1 has the characteristic active-site disulfide bond of the carbapenemase family of beta-lactamases and has a structure that is very similar to those of other known carbapenemases, including NMC-A, SME-1 and KPC-2. Most residues implicated in the catalytic mechanism of this class of enzyme are present in the GES-1 active site, including Ser70, which forms a covalent bond with the carbonyl C atom of the beta-lactam ring of the substrate during the formation of an acyl-enzyme intermediate, Glu166, which is implicated as both the acylation and deacylation base, and Lys73, which is also implicated as the acylation base. A water molecule crucial to catalysis is observed in an identical location as in other class A beta-lactamases, interacting with the side chains of Ser70 and Glu166. One important residue, Asn170, also normally a ligand for the hydrolytic water, is missing from the GES-1 active site. This residue is a glycine in GES-1 and the enzyme is unable to hydrolyze imipenem. This points to this residue as being critically important in the hydrolysis of this class of beta-lactam substrate. This is further supported by flexible-docking studies of imipenem with in silico-generated Gly170Asn and Gly170Ser mutant GES-1 enzymes designed to mimic the active sites of imipenem-hydrolyzing point mutants GES-2 and GES-5.

MeSH terms

  • Amino Acid Sequence
  • Binding Sites
  • Carbapenems / metabolism*
  • Crystallization
  • Crystallography, X-Ray
  • Klebsiella pneumoniae / enzymology
  • Models, Molecular
  • Molecular Sequence Data
  • Protein Structure, Tertiary
  • Sequence Homology, Amino Acid
  • beta-Lactamases / chemistry*
  • beta-Lactamases / classification
  • beta-Lactamases / metabolism*

Substances

  • Carbapenems
  • GES-1 beta-lactamase
  • beta-Lactamases