Enhanced direct ethanol production by cofactor optimization of cell surface-displayed xylose isomerase in yeast

Xylose isomerase (XylC) from Clostridium cellulovorans can simultaneously perform isomerization and fermentation of d-xylose, the main component of lignocellulosic biomass, and is an attractive candidate enzyme. In this study, we optimized a specified metal cation in a previously established Saccharomyces cerevisiae strain displaying XylC. We investigated the effect of each metal cation on the catalytic function of the XylC-displaying S. cerevisiae. Results showed that the divalent cobalt cations (Co²⁺ ) especially enhanced the activity by 46-fold. Co²⁺ also contributed to d-xylose fermentation, which resulted in improving ethanol yields and xylose consumption rates by 6.0- and 2.7-fold, respectively. Utility of the extracellular xylose isomerization system was exhibited in the presence of mixed sugar. XylC-displaying yeast showed the faster d-xylose uptake than the yeast producing XI intracellularly. Furthermore, direct xylan saccharification and fermentation was performed by unique yeast co-culture system. A xylan-degrading yeast strain was established by displaying two kinds of xylanases; endo-1,4-β-xylanase (Xyn11B) from Saccharophagus degradans, and β-xylosidase (XlnD) from Aspergillus niger. The yeast co-culture system enabled fine-tuning of the initial ratios of the displayed enzymes (Xyn11B:XlnD:XylC) by adjusting the inoculation ratios of Xylanases (Xyn11B and XlnD)-displaying yeast and XylC-displaying yeast. When the enzymes were inoculated at the ratio of 1:1:2 (1.39 × 10¹³ : 1.39 × 10¹³ : 2.78 × 10¹³ molecules), 6.0 g/L ethanol was produced from xylan. Thus, the cofactor optimization and the yeast co-culture system developed in this study could expand the prospect of biofuels production from lignocellulosic biomass. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1068-1076, 2017.