Abstract

Efficient xylose fermentation by yeast would improve the economical and sustainable nature of biofuels production from lignocellulosic biomass. However, the efficiency of xylose fermentation by the yeast Saccharomyces cerevisiae is suboptimal, especially in conversion yield, despite decades of research. Here, we present an improved performance of S. cerevisiae in xylose fermentation through systematic and evolutionary engineering approaches. The engineering of S. cerevisiae harboring xylose isomerase-based pathway significantly improved the xylose fermentation performance without the need for intensive downstream pathway engineering. This strain contained two integrated copies of a mutant xylose isomerase, gre3 and pho13 deletion and XKS1 and S. stipitis tal1 overexpression. This strain was subjected to rapid adaptive evolution to yield the final, evolved strain (SXA-R2P-E) which could efficiently convert xylose to ethanol with a yield of 0.45 g ethanol/g xylose, the highest yield reported to date. The xylose consumption and ethanol production rates, 0.98 g xylose g cell−1 h−1 and 0.44 g ethanol g cell−1 h−1, respectively, were also among the highest reported. During this process, the positive effect of a pho13 deletion was identified for a xylose isomerase-containing strain and resulted in up to an 8.2-fold increase in aerobic growth rate on xylose. Moreover, these results demonstrated that low inoculum size and the cell transfer at exponential phase was found to be the most effective adaptation strategy during a batch culture adaptation process. These results suggest that the xylose isomerase pathway should be the pathway of choice for efficient xylose fermentation in S. cerevisiae as it can outperform strains with the oxidoreductase pathway in terms of yield and ethanol production and xylose consumption rates. Consequently, the strain developed in this study could significantly improve the prospect of biofuels production from lignocellulosic biomass.

Highlights

  • Efficient xylose fermentation by yeast would improve the economical and sustainable nature of biofuels production from lignocellulosic biomass

  • Episomal expression of xylA3* combined with overexpression of the native XKS1 and tal1 from S. stipitis resulted in improved growth rates and xylose consumption rates [5]

  • The effect of pho13 deletion with a xylose isomerase pathway In an effort to further improve xylose consumption rates, we evaluated the impact of a pho13 deletion on the xylose isomerase pathway

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Summary

Introduction

Efficient xylose fermentation by yeast would improve the economical and sustainable nature of biofuels production from lignocellulosic biomass. The efficiency of xylose fermentation by the yeast Saccharomyces cerevisiae is suboptimal, especially in conversion yield, despite decades of research. Whole-cell evolutionary engineering has been routinely applied to these rationally engineered strains to boost xylose fermentation efficiency [16,17,18]. Despite this array of attempts, the efficiency (especially yield) of xylose fermentation still remains suboptimal to achieve the goal of economical and sustainable biofuel production from lignocellulosic biomass, especially when compared with glucose conversion

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