Development of a superior industrial yeast strain for consolidated bioprocessing of lignocellulosic biomass

Worldwide, there is rapidly increasing interest in the production of biofuels and bio-based chemicals from lignocellulosic waste streams. An essential requirement for such processes is the availability of a micro-organism that is able to utilize all sugars, including the major pentose sugar xylose, and has high tolerance to the inhibitory compounds present in lignocellulosic hydrolysates. In addition, since the cost of the enzymes needed for hydrolysis of the lignocellulosic biomass still represents a significant percentage of the cost of the final product, a microorganism that secretes lignocellulolytic enzymes to reduce this cost at least to some extent, would be very useful to ensure economic viability of bio-based production. In this project, we will improve xylose fermentation capacity, inhibitor and thermo-tolerance, as well as lignocellulolytic enzyme secretion in an industrial Saccharomyces cerevisiae yeast strain for second-generation production of bioethanol and which can be used as a chassis strain for production of bio-based chemicals. For that purpose, we will improve the stability of the xylose uptake carriers and engineer mutant alleles that confer higher inhibitor and higher thermo-tolerance. We will enrich the gene pool further by whole-genome transformation using DNA from inhibitor- and thermo-tolerant non-conventional yeast species. We will also perform evolutionary adaptation with alternating lignocellulosic substrate conditions to select for higher production of the different lignocellulolytic enzymes. Mutations causing negative side-effects will be eliminated while beneficial mutations will be maintained and enriched by genome shuffling. This will result in an industrial yeast strain that can ferment at 40°C all glucose and xylose sugar in undetoxified bagasse hydrolysate in less than 40h and with a savings of at least 20% of the required enzyme load for bagasse hydrolysis.