Biofuels And Chemicals Production From Renewable Raw-Materials. Exploiting yeasts diversity to bridge the gap between the proof-of-concept and industrial success

The success of the biorefinery concept will require efficient, robust and versatile cell factories. Currently, the major part of industrial microorganisms are used because of historical grounds, rather than being selected for a specific application. Additionally, demands for increased productivity, wider substrate range utilization, and production of nonconventional compounds lead to a great interest in further improving the currently used industrial workhorses (hosts) and the selection or development of strains with novel properties. The model yeast Saccharomyces cerevisiae is the main microorganism used for first generation ethanol production. When moving from first to second generation of production, one of the major obstacles for a viable development is the toxic effect of compounds released during the pre-treatment of lignocellulosic biomasses, which are the more sustainable feedstock utilized. In the first part of this work, two different approaches to improve S. cerevisiae tolerance to compounds deriving from biomass pre-treatment are described. Firstly, the effects of overexpressing genes encoding the transcription factor (YAP1) and the mitochondrial NADH-cytochrome b5 reductase (MCR1) was evaluated in an industrial xylose-consuming S. cerevisiae strain. During batch fermentation on undiluted and undetoxified spruce hydrolysate overexpression of either genes resulted in faster hexose catabolism. The second approach revealed that acetic acid tolerance of S. cerevisiae can be increased by engineering it to endogenously produce L-ascorbic acid (L-AA). In the second part of the work, since the currently used industrial yeasts represent only the tip of the proverbial iceberg of the genetic diversity present in nature, different non-saccharomyces yeasts were investigated for their potential industrial applications: Kluyveromyces marxianus (CBS 712), the oleaginous yeasts Rhodosporidium toruloides (DSM 4444), Lipomyces starkeyi (DSM 70295) and Cryptococcus curvatus (DSM 70022), Zygosacchromyces bailii and finally, Candida lignohabitans. Overall, the work performed resulted in the development of industrial S. cerevisiae strains with improved traits that can match the requirements of lignocellulosic hydrolysate fermentation. The work also contributed to a better understanding of the metabolism and physiology of different non-saccharomyces yeasts with a great industrial potential.