Pichia stipitis xylose reductase helps detoxifying lignocellulosic hydrolysate by reducing 5-hydroxymethyl-furfural (HMF)

Background: Pichia stipitis xylose reductase (Ps-XR) has been used to design Saccharomyces
cerevisiae strains that are able to ferment xylose. One example is the industrial S. cerevisiae xyloseconsuming
strain TMB3400, which was constructed by expression of P. stipitis xylose reductase and
xylitol dehydrogenase and overexpression of endogenous xylulose kinase in the industrial S.
cerevisiae strain USM21.
Results: In this study, we demonstrate that strain TMB3400 not only converts xylose, but also
displays higher tolerance to lignocellulosic hydrolysate during anaerobic batch fermentation as well
as 3 times higher in vitro HMF and furfural reduction activity than the control strain USM21. Using
laboratory strains producing various levels of Ps-XR, we confirm that Ps-XR is able to reduce HMF
both in vitro and in vivo. Ps-XR overexpression increases the in vivo HMF conversion rate by
approximately 20%, thereby improving yeast tolerance towards HMF. Further purification of Ps-XR
shows that HMF is a substrate inhibitor of the enzyme.
Conclusion: We demonstrate for the first time that xylose reductase is also able to reduce the
furaldehyde compounds that are present in undetoxified lignocellulosic hydrolysates. Possible
implications of this newly characterized activity of Ps-XR on lignocellulosic hydrolysate
fermentation are discussed.