Construction of Recombinant Yeast Expressing Thermo-StableEnzyme for Efficient Bioethanol Production from Woody Biomass

Xylitol dehydrogenase from Pichia stipitis (PsXDH) is one of the key enzymes for the bioethanol fermentation system from biomass sugars. Previously, the C4 mutant (S96C/S99C/Y102C) was constructed with enhanced thermostability by introduction of structural zinc. A high thermostability of PsXDH was obtained by subsequent site-directed mutagenesis of the structural zinc-binding loop. In this study, effects of thermostabilization of xylitol dehydrogenase (XDH) from Pichia stipitis on fermentation of xylose to ethanol were estimated using a recombinant Saccharomyces cerevisiae expressing together with a native xylose reductase (XR) from P. stipitis. The mutated XDH performed the similar enzyme properties in S. cerevisiae cells, compared those in vitro. The significant enhancement was found in Y-C4 strain, 51% decrease of unfavorable xylitol excretion with 19% increased ethanol production, when compared with the reference strain expressing the wild–type XDH.