Poster Presentation 2-33

Engineering Redox Cofactor Regeneration for Improved D-Xylose Fermentation in Yeast

Ritva Verho, John Londesborough, Merja Penttilä and Peter Richard*

VTT Biotechnology
PO Box 1500

02044 VTT, Finland

Phone: +358 9 456 7190

Fax: +358 9 455 2103

E-mail: Peter.Richard@vtt.fi

Pentose fermentation to ethanol with recombinant Saccharomyces cerevisiae is slow and has a low yield. A likely reason for this is that the catabolism through the corresponding fungal pathway creates an imbalance of redox cofactors. The redox neutral process requires NADPH and NAD⁺ which have to be regenerated in separate processes. To facilitate NADPH regeneration we expressed the recently discovered gene GDP1 coding for a fungal NADP⁺ dependent D-glyceraldehyde 3-phosphate dehydrogenase in a S. cerevisiae strain with the D-xylose pathway. The NADPH regeneration through an NADP-GAPDH is not linked to CO₂ production. The oxidative part of the pentose phosphate pathway is the main natural path for NADPH regeneration. However use of this pathway causes wasteful CO₂ production and creates a redox imbalance on the path of anaerobic D-xylose fermentation to ethanol because it does not regenerate NAD⁺. The deletion of ZWF1, to shut down the oxidative part of the pentose phosphate pathway, in combination with overexpression of GDP1 stimulated D-xylose fermentation with respect to rate and yield. Through genetic engineering of the redox reactions, the yeast strain was converted from a strain that mainly produced xylitol and CO₂ from D-xylose to a strain that produced mainly ethanol in anaerobic conditions.