Enhanced Xylitol Production in Engineered Saccharomyces Cerevisiae By Dual Utilization of NADPH and NADH Cofactors

Xylitol, a natural sweetener, can be produced by hydrogenation of xylose in hemicelluloses. In microbial processes, utilization of only NADPH cofactor limited xylitol biosynthesis not to reach an industrial level. To overcome this drawback, Saccharomyces cerevisiae D452-2 was engineered to express two types of a xylose reductase(XR) with either NADPH-dependence or NADH-preference. Engineered S. cerevisiae DWM expressing both the XRs exhibited 10% higher xylitol productivity than yeast strains expressing each XR in batch culture. Furthermore, the coexpression of S. cerevisiae ZWF1 and ACS1 genes in the DWM-ZWF1-ACS1 strain increased intracellular concentrations of NADPH and NADH, and improved maximum xylitol productivity by 25%. Finally, the optimized fed-batch fermentation of S. cerevisiae DWM-ZWF1-ACS1 resulted in 196.2 g/L xylitol concentration, 4.27 g/L-h productivity and almost the theoretical yield. Expression of the two types of XR utilizing both NADPH and NADH is a promising strategy to meet the industrial demands for microbial xylitol production.