Arabidopsis glyoxalase II contains a zinc/iron binuclear metal center thatis essential for substrate binding and catalysis

Glyoxalase II participates in the cellular detoxification of cytotoxic and mutagenic 2-oxoaldehydes. Because of its role in chemical detoxification, g lyoxalase II has been studied as a potential anti-cancer and/or antiprotozo al target; however, very little is known about the active site and reaction mechanism of this important enzyme. To characterize the active site and ki netic mechanism of the enzyme, a detailed mutational study of Arabidopsis g lyoxalase II was conducted. Data presented here demonstrate for the first t ime that the cytoplasmic form of Arabidopsis glyoxalase II contains an iron -zinc binuclear metal center that is essential for activity. Both metals pa rticipate in substrate binding, transition state stabilization, and the hyd rolysis reaction. Subtle alterations in the geometry and/or electrostatics of the binuclear center have profound effects on the activity of the enzyme . Additional residues important in substrate binding have also been identif ied. An overall reaction mechanism for glyoxalase II is proposed based on t he mutational and kinetic data from this study and crystallographic data on human glyoxalase II. Information presented here provides new insights into the active site and reaction mechanism of glyoxalase II that can be used f or the rational design of glyoxalase II inhibitors.