Glyoxalase II from Salmonella typhimurium: new insghts in metal ion selectivity

VALERIA A. CAMPOS BERMUDEZ, LEITE RIBEIRO, R. KROG, A.J. COSTA-FILHO, F.C. SONCINI, G. OLIVA, A.J. VILA

Montevideo-Uruguay

Congreso; 6th International Conference of Biological Physics ICBP 2007, 5th Southern Cone Biophysics Congress and 34th Annual Meeting of the Argentinean Biophysical Society; 2007

Sociedad Argentina de Biofisica

Glyoxalase II is a hydrolytic enzyme part of the glyoxalase system, responsible fordetoxifying several cytotoxic compounds employing glutathione. Glyoxalase II belongs tothe superfamily of metallo-β-lactamases, with a conserved motif able to bind up to twometal ions in their active sites, generally zinc. Instead, several eukaryotic glyoxalases IIhave been characterized with different ratios of iron, zinc and manganese ions. We haveexpressed a gene coding for a putative member of this enzyme superfamily fromSalmonella enterica serovar Typhimurium. Based on activity assays we found it to be aglyoxalase II which we named GloB. Recombinant GloB expressed in Escherichia coli waspurified with variable amounts of iron, zinc and manganese. All forms display similaractivities, as can be shown from protein expression in minimal medium supplemented withspecific metal ions. The crystal structure of GloB solved at 1.4 Å shows a protein fold andactive site similar to those of its eukaryotic homologues. NMR and EPR experiments alsoreveal a conserved electronic structure at the metal site. GloB is therefore able toaccommodate these different metal ions and to carry out the hydrolytic reaction withsimilar efficiencies in all cases. The metal promiscuity of this enzyme (in contrast to othermembers of the same superfamily) can be accounted for by the presence of a conservedAsp residue acting as a second-shell ligand that is expected to increase the hardness of themetal binding site, therefore favoring iron uptake in glyoxalases II. However, thepossibility of that this enzyme employs different metal ions during different stage of its lifecyclecannot be ruled out.