GALACTOSE EXPULSION DURING LACTOSE METABOLISM IN LACTOCOCCUS-LACTIS SUBSP CREMORIS FD1 DUE TO DEPHOSPHORYLATION OF INTRACELLULAR GALACTOSE 6-PHOSPHATE

In Lactococcus lactis subsp. cremoris FD1, galactose and lactose are b oth transported and phosphorylated by phosphotransferase systems. Lact ose 6-phosphate (lactose-6P) is hydrolyzed intracellularly to galactos e-6P and glucose. Glucose enters glycolysis as glucose-6P, whereas gal actose-6P is metabolized via the tagatose-6P pathway and enters glycol ysis at the tagatose diphosphate and fructose diphosphate pool. Galact ose would therefore be a gluconeogenic sugar in L. lactis subsp. cremo ris FD1, but since fructose 1,6-diphosphatase is not present in this s train, galactose cannot serve as an essential biomass precursor (gluco se-6P or fructose-6P) but only as an energy (ATP) source. Analysis of the growth energetics shows that transition from N limitation to limit ation by glucose-6P or fructose-6P gives rise to a very high growth-re lated ATP consumption (152 mmol of ATP per g of biomass) compared with the value in cultures which are not limited by glucose-6P or fructose -6P (15 to 50 mmol of ATP per g of biomass). During lactose metabolism , the galactose flux through the tagatose-6P pathway (r(max) = 1.2 h-1 ) is lower than the glucose flux through glycolysis (r(max) = 1.5 h-1) and intracellular galactose-6P is dephosphorylated; this is followed by expulsion of galactose. Expulsion of a metabolizable sugar has not been reported previously, and the specific rate of galactose expulsion is up to 0.61 g of galactose g of biomass -1 h-1 depending on the lac tose flux and the metabolic state of the bacteria. Galactose excreted during batch fermentation on lactose is reabsorbed and metabolized whe n lactose is depleted from the medium. In vitro incubation of galactos e-6P (50 mM) and permeabilized cells (8 g/liter) gives a supernatant c ontaining free galactose (50 mM) but no P(i) (less than 0.5 mM). No or ganic compound except the liberated galactose is present in sufficient concentration to bind the phosphate. Phosphate is quantitatively reco vered in the supernatant as P(i) by hydrolysis with alkaline phosphata se (EC 3.1.3.1), whereas inorganic pyrophosphatase (EC 3.6.1.1) cannot hydrolyze the compound. The results indicate that the unknown phospha te-containing compound might be polyphosphate.