Purification and characterization of 3-deoxyglucosone metabolizing enzyme from Bacillus sp.2.

A NADPH-dependent 3-DG metabolizing enzyme was isolated and purified to electrophoretic homogeneity from Bacillus sp.2 by combined consecutive treatment consisting of ammonium sulfate fractionation, Q Sepharose FF, Sephadex G-100 (I), Hydroxyapatite and Sephadex G-100 (II) column chromatographies. The specific activity of purified 3-DG metabolizing enzyme was 63.75 U/mg. The molecular weight of the enzyme was about 32 ku. 2-Oxoaldehyde compounds were found to be specifically good substrate for this reductase. The optimum pH of the enzyme activity was 6.2. The enzyme was stable in the pH range from 5 to 8 and in the temperature range from 25 degrees C to 30 degrees C. The K-m for 3-DG was 2.3 mmol/L. Suitable amount of EDTA, beta-mercaptoethanol and dithiothreitol enhanced the enzyme activity, hut the activity of the enzyme was partially lost by adding iodoacetic acid or N-ethylmaleimide.