Engineering a xylose fermenting yeast for lignocellulosic ethanol production

Abstract

Lignocellulosic ethanol is produced by yeast fermentation of lignocellulosic hydrolysates generated by chemical pretreatment and enzymatic hydrolysis of plant cell walls. The conversion of xylose into ethanol in hydrolysates containing microbial inhibitors is a major bottleneck in biofuel production. We identified sodium salts as the primary yeast inhibitors, and evolved a Saccharomyces cerevisiae strain overexpressing xylose catabolism genes in xylose or glucose-mixed medium containing sodium salts. The fully evolved yeast strain can efficiently convert xylose in the hydrolysates to ethanol on an industrial scale. We elucidated that the amplification of xylA, XKS1 and pentose phosphate pathway-related genes TAL1, RPE1, TKL1, RKI1, along with mutations in NFS1, TRK1, SSK1, PUF2 and IRA1, are responsible and sufficient for the effective xylose utilization in corn stover hydrolysates containing high sodium salts. Our evolved or reverse-engineered yeast strains enable industrial-scale production of lignocellulosic ethanol and the genetic foundation we uncovered can also facilitate transfer of the phenotype to yeast cell factories producing chemicals beyond ethanol.