GLYOXALASE-II FROM ARABIDOPSIS-THALIANA REQUIRES ZN(II) FOR CATALYTICACTIVITY

Cytosolic glyoxalase II from Arabidopsis thaliana, GLXZ-2, was overexp ressed and purified to homogeneity using Q-sepharose chromatography. M ALDI-TOF mass spectrometry studies indicated a molecular weight of 28 767 Da, Using steady-state kinetics studies, the purified enzyme exhib ited a K-m of 660 +/- 100 mu M and a k(cat) of 484 +/- 92 s(-1) at 37 degrees C, Metal analyses demonstrated that the enzyme binds 2.1 +/- 0 .5 moles of Zn(II) per monomer; the binding of ZnO is essential for en zyme viability and activity. Sequence comparison of glyoxalase II enzy mes from human, A. thaliana, and yeast and the metallo-beta-lactamases reveal that all metal binding Ligands of the metallo-beta-lactamases are conserved in glyoxalase Il enzymes, suggesting that all glyoxalase II enzymes are Zn(II) metalloenzymes. These results and their implica tions are discussed in Light of previous studies on glyoxalase II, and an active site for the glyoxalase II enzymes is proposed. (C) 1997 Fe deration of European Biochemical Societies.