Glucose repression of the TCA cycle in Saccharomyces cerevisiae was investigated under different environmental conditions by using (13)C-tracer experiments. Real-time quantification of the volatile metabolites ethanol and CO(2) allowed accurate carbon balancing. In all experiments with the wild-type, a strong correlation between the growth and glucose uptake rates was observed, indicating a constant yield of biomass. In contrast, glycerol and acetate production rates were less dependent on the rate of glucose uptake, but were more affected by environmental conditions. The glycerol production rate (2.9 mmol/g/h) was highest during growth in high osmolarity medium, while the highest acetate production rate of 2.1 mmol/g/h was observed in alkaline medium of pH 6.9. S. cerevisiae had, under standard growth conditions (25 g/L glucose, pH 5.0, 30 degrees C), low fluxes through the pentose phosphate pathway and the TCA cycle. A significant increase in TCA cycle activity from 0.03 mmol/g/h to about 1.7 mmol/g/h was observed when S. cerevisiae grew slower as a result of environmental perturbations, including unfavourable pH values and sodium chloride stress. Compared to experiments with high glucose uptake rates, the ratio of CO(2) to ethanol increased more than 50%, indicating an increase in flux through the TCA cycle. Although glycolysis and the ethanol production pathway still exhibited the highest fluxes, the net flux through the TCA cycle increased significantly with decreasing glucose uptake rates. Results from experiments with single gene deletion mutants partially impaired of glucose repression (hxk2, grr1) indicate that the rate of glucose uptake correlate to this increase in TCA cycle flux. These findings are discussed in the context of regulation of glucose repression.
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