Effect of enhanced xylose reductase activity on xylose consumption and product distribution in xylose-fermenting recombinant Saccharomyces cerevisiae

Abstract

Recombinant Saccharomyces cerevisiae TMB3001, harboring the Pichia stipitis genes XYL1 and XYL2 (xylose reductase and xylitol dehydrogenase, respectively) and the endogenous XKS1(xylulokinase), can convert xylose to ethanol. About 30% of the consumed xylose, however, is excreted as xylitol. Enhanced ethanol yield has previously been achieved by disrupting the ZWF1 gene, encoding glucose-6- phosphate dehydrogenase, but at the expense of the xylose consumption. This is probably the result of reduced NADPH-mediated xylose reduction. In the present study, we increased the xylose reductase (XR) activity 4^19 times in both TMB3001 and the ZWF1-disrupted strain TMB3255. The xylose consumption rate increased by 70% in TMB3001 under oxygen-limited conditions. In the ZWF1-disrupted background, the increase in XR activity fully restored the xylose consumption rate. Maximal specific growth rates on glucose were lower in the ZWF1-disrupted strains, and the increased XR activity also negatively affected the growth rate in these strains. Addition of methionine resulted in 70% and 50% enhanced maximal specific growth rates for TMB3255 (zwf1v) and TMB3261 (PGK1-XYL1, zwf1v), respectively. Enhanced XR activity did not have any negative effect on the maximal specific growth rate in the control strain. Enhanced glycerol yields were observed in the high-XR-activity strains. These are suggested to result from the observed reductase activity of the purified XR for dihydroxyacetone phosphate.