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An enzyme catalyzing the reduction of methylglyoxal was isolated from Saccharomyces wrevisiae and its enzymatic properties were analyzed. The enzyme, specifically eluted from a blue-dextran -Sepharose CL-6B column by the substrate, methylglyoxal, was homogeneous on polyacrylamide gel electrophoresis. The enzyme consisted of single polypeptide chain with a relative molecular mass of 43000. The enzyme was glycoprotein and contained 6.6% carbohydrate. NADPH was specifically required for activity and the K, for NADPH was 2.0 x lo-' M. The enzyme was active on various glyoxals such as glyoxal, methylglyoxal ( K , = 5.88 mM) and phenylglyoxal ( K , = 1.54 mM). The reaction catalyzed by the enzyme was virtually irreversible. The aclivity was inhibited by sulfhydryl agents and activated by reducing agents such as glutathione. Intermediates in glycolysis, nucleosides, nucleotides, polyamines and various metal ions showed little inhibitory or activating effects on enzyme activity. Tricarboxylic acids showed a slight inhibitory effect. The activity of the enzyme was strongly inhibited by anionic detergents. The enzyme was rapidly inactivated by incubating with the substrates probably because of the non-enzymatic interaction between glyoxals and NH2 groups in arginine residues in the enzyme. NADP, one of the reaction products, also inhibited the enzyme activity and the Ki for NADP was about 0.07 mM.We tentatively designated the enzyme methylglyoxal reductase.

NAD-dependent lactaldehyde dehydrogenase, catalyzing an oxidation of lactaldehyde to lactate, was purified approximately 70-fold from cell extracts of Saccharomyces cerevisiae with a 28% yield of activity. The enzyme was homogeneous on polyacrylamidc gel electrophoresis. The relative molecular mass of the enzyme was estimated to be 40000 on Sephadex G-150 column chromatography and on sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The enzyme was most active at pH 6.5, 60 -C and specifically oxidized L-lactaldehyde to L-lactate in the presence of NAD. The K, values for L-lactaldehyde and NAD were 10 mM and 2.9 mM, respectively. The purest enzyme was extremely unstable and almost completely inactivated during storage at -20 C, pH 7.5. For the reactivation of the enzyme, halide ions such as CI-, I and Br-were required.The metabolism of lactaldehyde in yeast cells has been studied by Neuberg and Vercellone [I] and then by Sandman and Miller [2]. Their investigations were designcd to study only the metabolic relationship between lactaldehyde, acetol (1 -hydroxy-2-propanone), 1,2-propane-diol and lactate. The data obtained by them indicated that the lactaldehyde was converted to 1 ,2-propane-diol, lactate and acetol, thc acetol probably being derived from 1,2-propane-dioI [ 11. However, no data were given for the nature of the enzymes involved in the metabolism of these compounds.Recently, we found that yeast cells of Saccharomyces wrrvisiae could convert methylglyoxal to lactaldehyde by the action of NADPH-dependent methylglyoxal reductase and the reaction catalyzed by the reductase was virtually irreversible [3]. Further enzymatic oxidation of lactaldehyde is, therefore, considered to result in either of two products, 1,2-propane-diol and lactate. To elucidate the fate of lactaldehyde formed from methylglyoxal, the enzyme catalyzing the oxidation of lactaldehyde to lactate was sought, purified and its properties were characterized. MATERIALS AND METHODS Chemicalswere cultured in a nutrient medium (1.0% yeast extract, 2.0% peptone, 2.0% glucose, pH 5.0; 1.5 1 medium in a 2-1 Sakaguchi flask) at 30°C for 24 h with reciprocation. Cells (55 g as wet weight from 10 1 of culture) after centrifugation were washed once with 0.85% saline solution, resuspended in 10 mM Tris/HCI buffer (pH 7.5) containing 0.1 mM methylphenylsulfonyl fluoride (PhMeS02F) [Tris/ PhMeSO,F]. The cell suspension was homogenized on a Dyno-Mill for 4 min at 0' C. The supernatant after centrifugation at 8000 rpm was used for the purification of lactaldehyde dehydrogenase. Assa,yfor lactaldehyde dehydrogenaseThe reaction mixture (3.0 ml) contained 4.3 mM L-lactaldehyde, 5.0 mM NAD, 50 mM Tris/HCl buffer (pH 6.5) and enzyme. The reaction was initiated by the addition of enzyme and an increase in absorbance at 340 nm at 25°C was recorded on a Hitachi spectrophotometer model 100-50. Lactaldehyde concentration was determined by bisulfate uptake [5]. One unit of enzyme activity was defined as the amount of enzyme catalyzing the formation of 1 .O...