Impact of complex inhibitors in straw hydrolysates on xylose-fermenting Saccharomyces cerevisiae and inhibitor-tolerant strain construction.

Enhancing the ability of Saccharomyces cerevisiae strains to ferment mixed sugars under complex inhibitors is crucial for boosting the economic feasibility of cellulosic ethanol production. Using comparative transcriptomics, this study explored the impact of combined inhibitors (acetic acid, formic acid, furfural, and vanillin) on the xylose-fermenting S. cerevisiae strain s6 during mixed-sugar fermentation. The results revealed significant disruptions in carbon metabolism, amino acid metabolism, nucleic acid metabolism, energy metabolism, and signal transduction pathways, suggesting that cells may redirect energy from biosynthetic processes to combat inhibitor stress. Based on these findings, eight genes, including five transcription factors and three functional genes, were selected for combinatorial overexpression. Fermentation assessments in both inhibitors-added artificial media and real straw hydrolysates confirmed improved inhibitor tolerance, particularly in strains with optimized gene combinations. Strains s6H3F7T6 (overexpressing HAA1, FDH1, and TYE7) and s6H3F7 (overexpressing HAA1 and FDH1) exhibited significantly enhanced xylose consumption rates, increasing by 137.48 % and 133.43 %, respectively, along with improved ethanol production of 19.54 % and 10.37 % in artificial media containing a mixture of four inhibitors. Notably, s6H3F7 and s6H3F7T6 also demonstrated substantial improvements in two types of straw hydrolysates containing a multitude of inhibitors. This study validates the effectiveness of simultaneous multi-gene regulation, providing robust resistant strains with substantial promise for industrial applications.