P153: Engineering of baker’s yeast for production of bioethanol from mixed sugars

Bioethanol is utilized as the gasoline additive, which provides a transportation fuel for current use. Industrial production of bioethanol has been conventionally conducted by Baker’s yeast Saccharomyces cerevisiae using sugar-containing feedstock. However, developing a sustainable production process of bioethanol from renewable resources now becomes a focus of research. Lignocellulose is the most abundant resource and hydrolyzed to glucose and xylose. Unfortunately, native S. cerevisiae strain is unable to utilize xylose. In this study, a genetic toolbox based on a recyclable DNA cassette was developed for site-specific insertion of an artificial promoter and of a heterologous gene. By using this genetic toolbox, the heterologous genes comprising the xylose-utilizing pathway were integrated into S. cerevisiae genome. Moreover, the endogenous XKS1 gene and genes involved in the pentose phosphate pathway were all fused with an artificial promoter to enhance their expression activity. Finally, xylose metabolism of the yeast strain was further improved by adaptive evolution. After evolution for 6 months, one evolved strain was scored and shown to effectively consume xylose. With glucose (10 g/L) and xylose (40 g/L), the evolved strain was able to consume all the sugars and produce 20 g/L ethanol, which accounts for a conversion yield of 0.4 g/g sugars.