Biochemical basis for glucose-induced inhibition of malolactic fermentation in Leuconostoc oenos

The sugar-induced inhibition of malolactic fermentation in cell suspensions of Leuconostoc oenos, recently reclassified as Oenococcus oeni (L. M. T. Dicks, F. Dellaglio, and M. D. Collins, Int. J. Syst. Bacteriol. 45:395-397, 1995) was investigated by in vivo and in vitro nuclear magnetic resonance (NMR) spectroscopy and manometric techniques. At 2 mM, glucose inhibited malolactic fermentation by 50%, and at 5 mM or higher it caused a maximum inhibitory effect of ca. 70%. Galactose, trehalose, maltose, and mannose caused inhibitory effects similar to that observed with glucose, but ribose and 2-deoxyglucose did not affect the rate of malolactic activity. The addition of fructose or citrate completely relieved the glucose-induced inhibition. Glucose was not catabolized by permeabilized cells, and inhibition of malolactic fermentation was not observed under these conditions. P-31 NMR analysis of perchloric acid extracts of cells obtained during glucose-malate cometabolism showed high intracellular concentrations of glucose-6-phosphate, 6-phosphogluconate, and glycerol-3-phosphate. Glucose-6-phosphate, 6-phosphogluconate, and NAD(P)H inhibited the malolactic activity in permeabilized cells or cell extracts, whereas NADP(+) had no inhibitory effect. The purified malolactic enzyme was strongly inhibited by NADH, whereas all the other above-mentioned metabolites exerted no inhibitory effect, showing that NADH was responsible for the inhibition of malolactic activity in vivo. The concentration of NADH required to inhibit the activity of the malolactic enzyme by 50% was ca. 25 mu M. The data provide a coherent biochemical basis to understand the glucose-induced inhibition of malolactic fermentation in L. oenos.