EBI Personnel Directory Dueber, John
In order to tap xylose, the major constituent of hemicellulose and an important step in sustainable biofuels, this project team is working to improve the efficiency of xylose catabolism in Saccharomyces cerevisiae. Enzyme expression is being optimized by systematically screening a library of promoters driving all enzymes in the pathway predicted to influence pathway flux, and bottlenecks at the protein level are being addressed by designing synthetic scaffolds to co-localize enzymes for increased enzymatic efficiency and to limit intermediate loss.
Currently, pectin-rich biomass, such as sugar beet pulp, apple pomace and citrus peels are considered low value waste streams, but these feedstocks can be inexpensively broken down into their component monosaccharides, primarily galacturonic acid (GalA), arabinose, and galactose. The generation of yeast strains with the ability to co-ferment these sugars would allow current waste streams to be converted into a sustainable biofuel. Our specific aims are the following: 1) determine enzyme and metabolite abundances in expression optimized strains enriched in both aerobic and anaerobic growth conditions with proteomics and metabolic flux analysis and use as an improved starting point for strain adaptations. 2) apply our combinatorial expression engineering strategy towards the utilization of galacturonic acid, the primary monosaccharide in pectic polysaccharides. 3) engineer a S.cerevisiae strain capable of co-utilizing galacturonic acid and arabinose.
A continuous fermentation process in S. cerevisiae would require simultaneous import of the major monosaccharides in lignocellulosic hydrolysates, namely glucose and xylose. Currently, both sugars are imported through the hexose transporters but these have a 10 to 100-fold preference for glucose over xylose. Recent directed evolution efforts by other groups have successfully made transporters that only import xylose, but glucose still inhibits xylose transport.