Stoichiometric network constraints on xylose metabolism by recombinant Saccharomyces cerevisiae.
Metabolic pathway engineering is constrained by the thermodynamic and stoichiometric feasibility of enzymatic activities of introduced genes. Engineering of xylose metabolism in Saccharomyces cerevisiae has focused on introducing genes for the initial xylose assimilation steps from Pichia stipitis, a xylose-fermenting yeast, into S. cerevisiae, a yeast traditionally used in ethanol production from hexose. However, recombinant S. cerevisiae created in several laboratories have used xylose oxidatively rather than in the fermentative manner that this yeast metabolizes glucose. To understand the differences between glucose and engineered xylose metabolic networks, we performed a flux balance analysis (FBA) and calculated extreme pathways using a stoichiometric model that describes the biochemistry of yeast cell growth. FBA predicted that the ethanol yield from xylose exhibits a maximum under oxygen-limited conditions, and a fermentation experiment confirmed this finding. Fermentation results were largely consistent with in silico phenotypes based on calculated extreme pathways, which displayed several phases of metabolic phenotype with respect to oxygen availability from anaerobic to aerobic conditions. However, in contrast to the model prediction, xylitol production continued even after the optimum aeration level for ethanol production was attained. These results suggest that oxygen (or some other electron accepting system) is required to resolve the redox imbalance caused by cofactor difference between xylose reductase and xylitol dehydrogenase, and that other factors limit glycolytic flux when xylose is the sole carbon source.
Jin
Yong-Su
YS
Department of Food Science, University of Wisconsin-Madison, 1605 Linden Drive, 53706, USA.
Jeffries
Thomas W
TW
D049109:Cell Proliferation; D003198:Computer Simulation; D000431:Ethanol / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology; D015966:Gene Expression Regulation, Fungal / Q000502:physiology; D008954:Models, Biological; D010100:Oxygen / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000502:physiology*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D014994:Xylose / Q000378:metabolism*
D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D000431:Ethanol; D014994:Xylose; D010100:Oxygen