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Background  Lignocellulosic biorefinery offers economical and sustainable production of fuels and chemicals.  Saccharomyces cerevisiae , a promising industrial host for biorefinery, has been intensively developed to expand its product profile. However, the sequential and slow conversion of xylose into target products remains one of the main challenges for realizing efficient industrial lignocellulosic biorefinery.    Results  In this study, we developed a powerful mixed-sugar co-fermenting strain of  S. cerevisiae , XUSEA, with improved xylose conversion capacity during simultaneous glucose/xylose co-fermentation. To reinforce xylose catabolism, the overexpression target in the pentose phosphate pathway was selected using a DNA assembler method and overexpressed increasing xylose consumption and ethanol production by twofold. The performance of the newly engineered strain with improved xylose catabolism was further boosted by elevating fermentation temperature and thus significantly reduced the co-fermentation time by half. Through combined efforts of reinforcing the pathway of xylose catabolism and elevating the fermentation temperature, XUSEA achieved simultaneous co-fermentation of lignocellulosic hydrolysates, composed of 39.6 g L −1  glucose and 23.1 g L −1  xylose, within 24 h producing 30.1 g L −1  ethanol with a yield of 0.48 g g −1 .    Conclusions  Owing to its superior co-fermentation performance and ability for further engineering, XUSEA has potential as a platform in a lignocellulosic biorefinery toward realizing a more economical and sustainable process for large-scale bioethanol production.

Improved simultaneous co-fermentation of glucose and xylose by Saccharomyces cerevisiae for efficient lignocellulosic biorefinery