Ping-Wei Ho

• In the past decade, the use of biodiesel as a fuel was considered as an alternative to reduce reliance on fossil resources and the resulting greenhouse effect. During biodiesel production, glycerol is generated as a by-product in excess. The economy of biodiesel industry is thus expected to benefit from the valorization of glycerol. One of the valorization avenues is employing glycerol as a substrate in microbial bioprocesses. The yeast Saccharomyces cerevisiae is a popular cell-factory in industrial biotechnology and a suitable chassis in synthetic biology. This status is the result of many factors, such as the vast knowledge available regarding the physiology and genetics of this species, the intense experience with this organism in industrial fermentations, S. cerevisiae’s robustness under process conditions, and the ease to genetically engineer this microorganism. Using glycerol as a carbon source for growing S. cerevisiae would bring about several assets for yeast-based bioprocesses. On one hand, glycerol does not trigger Crabtree effect in S. cerevisiae, which is in favor of bioprocesses aiming at high biomass production or biomass-associated products. On the other hand, glycerol is more reduced than sugars, which would be an advantage for the production of small-molecule compounds with a high degree of reduction. One of the main objectives of this thesis was to better understand glycerol utilization in S. cerevisiae and identify the underlying determinants for the better growth of certain strains on glycerol. The major focus was set on improving strains of the popular CEN.PK strain family. Another goal of the thesis was to provide thoroughly characterized S. cerevisiae promoters for the given cultivation conditions. Such information is required for achieving optimal metabolic engineering outcomes if glycerol is to be used as the sole carbon source.