Biochemical and structural characterization of Salmonella typhimurium glyoxalase II: New insights in metal ion selectivity

CAMPOS BERMUDEZ VA, RIBEIRO LEITE N, KROG R, COSTA-FILHO AJ , SONCINI FC , OLIVA G AND VILA AJ

BIOCHEMISTRY

American Chemical Society

Lugar: Washington; Año: 2007 vol. 46 p. 11069 - 11079

0006-2960

Glyoxalase II is a hydrolytic enzyme part of the glyoxalase system, responsible for detoxifying several cytotoxic compounds employing glutathione. Glyoxalase II belongs to the superfamily of metallo-beta-lactamases, with a conserved motif able to bind up to two metal ions in their active sites, generally zinc. Instead, several eukaryotic glyoxalases II have been characterized with different ratios of iron, zinc and manganese ions. We have expressed a gene coding for a putative member of this enzyme superfamily from Salmonella typhimurium that we demonstrate, based on its activity, to be a glyoxalase II, named GloB. Recombinant GloB expressed in Escherichia coli was purified with variable amounts of iron, zinc and manganese. All forms display similar activities, as can be shown from protein expression in minimal medium supplemented with specific metal ions. The crystal structure of GloB solved at 1.4 Å shows a protein fold and active site similar to those of its eukaryotic homologues. NMR and EPR experiments also reveal a conserved electronic structure at the metal site. GloB is therefore able to accommodate these different metal ions and to carry out the hydrolytic reaction with similar efficiencies in all cases. The metal promiscuity of this enzyme (in contrast to other members of the same superfamily) can be accounted for by the presence of a conserved Asp residue acting as a second-shell ligand that is expected to increase the hardness of the metal binding site, therefore favoring iron uptake in glyoxalases II.