Poster Presentation 2-14

Ethanol Formation from Xylose is Improved by Lowering the Oxidative Pentose-phosphate Pathway Flux

Marie Jeppsson, Björn Johansson, Bärbel Hahn-Hägerdal and Marie F. Gorwa-Grauslund

Applied Microbiology, P.O. Box 124, Chemical Center, 221 00 Lund, Sweden

Telephone:  (46) 46-222-8328; Fax:  (46) 46-222-4203; E-mail:  Marie.Jeppsson@tmb.lth.se

Saccharomyces cerevisiae efficiently ferments the hexose sugars of lignocellulosic hydrolysates to fuel ethanol.  However, it is unable to utilize xylose, the major pentose sugar of hardwood. Therefore, recombinant S. cerevisiae expressing the XYL1 and XYL2 genes from Pichia stipitis and overexpressing the native XKS1 gene have been constructed (Fig. 1).  XYL1 and XYL2 encode the NAD(P)H-dependent xylose reductase (Rizzi et al. 1988) and the NAD⁺-dependent xylitol dehydrogenase (Rizzi et al. 1989), respectively.  XKS1 encodes the ATP-consuming xylulokinase.  Anaerobic xylose fermentation has been demonstrated; however, a low ethanol yield was obtained, mainly because of xylitol formation (Eliasson et al. 2000).

We have shown that a disruption of oxidative pentose phosphate pathway (PPP) genes results in decreased xylitol- and increased ethanol yield, up to 0.41 g ethanol (g xylose)^-1 (Jeppsson et al. 2001).  This is, to the best of our knowledge, the highest ethanol yield ever obtained for recombinant xylose-utilising S. cerevisiae.  Since this strain had a low specific xylose consumption rate, we are working on strategies to enhance the rate of the strain by 1) increasing the expression of XR, and 2) allowing a small flux through the oxidative PPP.  Results from these experiments are presented. 

Figure 1. The xylose pathway which has been introduced in S. cerevisiae: Xylose reductase and xylitol dehydrogenase from Pichia stipitis, and native xylulokinase.   

Eliasson, A., C. Christensson, C. F. Wahlbom and B. Hahn-Hägerdal. (2000). “Anaerobic xylose fermentation by recombinant Saccharomyces cerevisiae carrying XYL1, XYL2, and XKS1 in mineral medium chemostat cultures.” Appl Environ Microbiol 66(8): 3381-6.

Jeppsson, M., B. Johansson, B. Hahn-Hägerdal, and M. F. Gorwa-Grauslund. (2001). “Reduced oxidative pentose phosphate pathway flux in recobinant Saccharomyces cerevisiae strains improves ethanol formation from xylose.” Submitted.

Rizzi, M., P. Erlemann, N.-A. Bui-Thanh, and H. Dellweg. (1988). “Xylose fermentation by yeast.  4. Purification and kinetic studies of xylose reductase from Pichia stipitis.” Appl Microbiol Biotechnol 29: 148-154.

Rizzi, M., P. Harwart, P. Ehrlemann, N. A. Bui-Thanh, and H. Dellweg. (1989). “Purification and properties of the NAD+ -xylitol-dehydrogenase from the yeast Pichia stipitis.” J Ferment Bioeng 67: 20-24.