Role of Hexose Transport in Control of Glycolytic Flux in Saccharomyces cerevisiae

Abstract

The yeast Saccharomyces cerevisiae predominantly ferments glucose to ethanol at high external glucose concentrations, irrespective of the presence of oxygen. In contrast, at low external glucose concentrations and in the presence of oxygen, as in a glucose-limited chemostat, no ethanol is produced. The importance of the external glucose concentration suggests a central role for the affinity and maximal transport rates of yeast's glucose transporters in the control of ethanol production. Here we present a series of strains producing functional chimeras between the hexose transporters Hxt1 and Hxt7, each of which has distinct glucose transport characteristics. The strains display a range of decreasing glycolytic rates resulting in a proportional decrease in ethanol production. Using these strains, we show for the first time that at high glucose levels, the glucose uptake capacity of wild-type S. cerevisiae does not control glycolytic flux during exponential batch growth. In contrast, our chimeric Hxt transporters control the rate of glycolysis to a high degree. Strains whose glucose uptake is mediated by these chimeric transporters will undoubtedly provide a powerful tool with which to examine in detail the mechanism underlying the switch between fermentation and respiration in S. cerevisiae and will provide new tools for the control of industrial fermentations.

Correlation between TCA cycle flux and glucose uptake rate during respiro-fermentative growth of Saccharomyces cerevisiae

“…Elbing et al (2004) reported the glucose uptake rate as the determinant of respiratory metabolism in S. cerevisiae. The use of chimeric hexose transporters allowed for the reduction of the maximal glucose uptake rate, leading to the absence of ethanol production at low uptake rates (below 5 mmol g 21 h…”

Progress in Metabolic Engineering of Saccharomyces cerevisiae

“…The absence of ethanol formation is particularly critical when S. cerevisiae is used for the production of heterologous proteins. A promising approach to obtaining yeast with a fully respiratory phenotype in sugar-containing industrial media such as molasses has been published by Elbing et al (82). They generated a yeast strain which is defective in the most important hexose transporter genes (HXT1 to -7) and expresses a chimeric hexose transporter.…”

“…It was mentioned in the previous section that recent attempts to abolish the Crabtree effect in yeast could be a novel avenue to reducing ethanol in beverages. Based on the approach of Elbing et al (82), a non-ethanol-producing wine yeast strain was developed by modifying hexose transporters (117). Moreover, introducing heterologous enzymes to increase NADH oxidation has been shown to reduce overflow metabolism, i.e., ethanol formation, in a wine yeast strain (121).…”

Engineering of a Novel Saccharomyces cerevisiae Wine Strain with a Respiratory Phenotype at High External Glucose Concentrations

“…In another recent report (8), we presented a series of strains expressing functional chimeras composed to different degrees of the low-affinity (K m , 100 mM) and high-affinity (K m , 1 to 2 mM) transporters Hxt1 and Hxt7, respectively (28); one of the strains was KOY.TM6*P (27). Sugar uptake analysis data showed that the Tm6* protein in the KOY.TM6*P strain proved to be a high-affinity hexose transporter (27).…”