Laboratory of Chemical Biotechnology, TU Dortmund, 44221 Dortmund, Germany.
Microbiology (Reading). 2009 Dec;155(Pt 12):3827-3837. doi: 10.1099/mic.0.030213-0. Epub 2009 Aug 14.
Glucose repression of the tricarboxylic acid (TCA) cycle in Saccharomyces cerevisiae was investigated under different environmental conditions 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 rates of growth and glucose uptake 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 affected by environmental conditions. The glycerol production rate was highest during growth in high-osmolarity medium (2.9 mmol g(-1) h(-1)), while the highest acetate production rate of 2.1 mmol g(-1) h(-1) was observed in alkaline medium of pH 6.9. Under standard growth conditions (25 g glucose l(-1) , pH 5.0, 30 degrees C) S. cerevisiae had low fluxes through the pentose phosphate pathway and the TCA cycle. A significant increase in TCA cycle activity from 0.03 mmol g(-1) h(-1) to about 1.7 mmol g(-1) h(-1) was observed when S. cerevisiae grew more slowly 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 in glucose repression (hxk2, grr1) indicated that the rate of glucose uptake correlates with this increase in TCA cycle flux. These findings are discussed in the context of regulation of glucose repression.
采用（13）C 示踪实验，在不同环境条件下研究了酿酒酵母三羧酸（TCA）循环的葡萄糖抑制作用。实时定量分析挥发性代谢物乙醇和 CO（2），实现了准确的碳平衡。在所有野生型实验中，都观察到生长速度和葡萄糖摄取率之间存在很强的相关性，表明生物质的产量保持恒定。相比之下，甘油和乙酸的产生速率与葡萄糖摄取率的关系较小，但受环境条件的影响。在高渗透压介质（2.9 mmol g（-1）h（-1））中生长时，甘油的产生速率最高，而在 pH 值为 6.9 的碱性介质中，乙酸的产生速率最高，达到 2.1 mmol g（-1）h（-1）。在标准生长条件（25 g 葡萄糖 l（-1），pH 值 5.0，30 摄氏度）下，酿酒酵母的戊糖磷酸途径和 TCA 循环通量较低。当由于环境干扰，包括不利的 pH 值和氯化钠胁迫，导致酿酒酵母生长缓慢时，TCA 循环的活性从 0.03 mmol g（-1）h（-1）显著增加到约 1.7 mmol g（-1）h（-1）。与葡萄糖摄取率较高的实验相比，CO（2）与乙醇的比值增加了 50%以上，表明 TCA 循环通量增加。尽管糖酵解和乙醇生产途径仍显示出最高的通量，但随着葡萄糖摄取率的降低，TCA 循环的净通量显著增加。用葡萄糖抑制作用部分受损的单基因缺失突变体（hxk2、grr1）进行实验的结果表明，葡萄糖摄取率与 TCA 循环通量的增加相关。这些发现将在葡萄糖抑制作用的调节背景下进行讨论。
