Kinetic modeling and sensitivity analysis for higher ethanol production in self-cloning xylose-using Saccharomyces cerevisiae

We constructed a xylose-utilizing Saccharomyces cerevisiae strain using endogenous xylose-assimilating genes (strain K7-XYL). Such self-cloning yeast is expected to make a great contribution to cost reduction of ethanol production processes. However, it is difficult to modify self-cloning yeast for optimal performance because the available gene source is limited. To improve the ethanol productivity of our self-cloning yeast, a kinetic model of ethanol production was constructed and sensitivity analysis was performed. Alcohol dehydrogenase (ADH1) was identified as a metabolic bottleneck reaction in the ethanol production pathway. An ADH1 overexpression strain (K7-XYL-ADH1) was constructed and evaluated in YP (yeast extract 10 g/L, peptone 20 g/L) medium containing 50 g/L xylose as the sole carbon source. Strain K7-XYL-ADH1 showed higher ethanol productivity (13.8 g/L) than strain K7-XYL (12.5 g/L). Then, K7-XYL-ADH1 was evaluated in YP medium containing 80 g/L glucose and 50 g/L xylose; however, the ethanol productivity did not change relative to that of K7-XYL (K7-XYL 46.3 g/L, K7-XYL-ADH1 45.9 g/L). We presumed that due to the presence of glucose, the internal redox balance of the cells had changed. On culturing in an aerated 5-L jar fermentor to change the internal redox balance of cells, strain K7-XYL-ADH1 showed higher ethanol productivity than K7-XYL (K7-XYL 45.0 g/L, K7-XYL-ADH1 49.4 g/L). Our results confirmed that ADH1 was a metabolic bottleneck in the ethanol production pathway. By eliminating the bottleneck, self-cloning yeast showed almost the same ethanol productivity as genetically modified yeast.