Overexpression of LAS21 in Cellulase-Displaying Saccharomyces cerevisiae for High-Yield Ethanol Production from Pretreated Sugarcane Bagasse

Abstract

The valorization of lignocellulosic feedstocks into biofuels and biochemicals has received much attention due to its environmental friendliness and sustainability. However, engineering an ideal microorganism that can both produce sufficient cellulases and ferment ethanol is highly challenging. In this study, we have tested seven different genes that are involved in glycosylphosphatidylinositol (GPI) biosynthesis and remodeling for the improvement of cellulase activity tethered on the S. cerevisiae cell surface. It was found that the overexpression of LAS21 can improve β-glucosidase activity by 48.8% compared to the original strain. Then, the three cellulase genes (cellobiohydrolase, endoglucanase, and β-glucosidase) and the LAS21 gene were co-introduced into a diploid thermotolerant S. cerevisiae strain by a multiple-round transformation approach, resulting in the cellulolytic ECBLCCE5 strain. Further optimization of the bioprocess parameters was found to enhance the ethanol yield of the ECBLCCE5 strain. Scaling up the valorization of pretreated sugarcane bagasses in a 1 L bioreactor resulted in a maximum ethanol concentration of 28.0 g/L (86.5% of theoretical yield). Our study provides a promising way to improve the economic viability of second-generation ethanol production. Moreover, the engineering of genes involved in GPI biosynthesis and remodeling can be applied to other yeast cell surface display applications.