Modular engineering of synthetic glycolytic pathways in Saccharomyces cerevisiae

Already for millennia, microbial fermentation is used for the production of dairy products, alcoholic beverages and bread. In the last decades, the field of biotechnology has tremendously expanded and nowadays, a wide range of compounds ranging from biofuels to chemicals and pharmaceuticals is produced using microbial cell factories. The development of genetic engineering tools has greatly contributed to this rapid development. Catalysing the conversion of renewable carbohydrate feedstocks into fuels and chemicals, microbial cell factories offer a sustainable alternative to fossil resources-based production, and thereby contribute to reduce greenhouse gas emissions. The yeast Saccharomyces cerevisiae plays an important role in industrial biotechnology. Its popularity for applied research and industrial production can be attributed to several factors as its fast fermentative metabolism, its tolerance to low pH, high sugar and alcohol concentrations and its genetic tractability. S. cerevisiae possesses one of the best furbished molecular toolboxes, which makes it possible to assemble complex heterologous pathways, as was recently illustrated by the successful biosynthesis of opioids in yeast. Despite this great progress, extensive genetic remodelling of native pathways remains challenging. This can largely be explained by the high genetic redundancy present in the yeast genome, in which multiple genes encode proteins with redundant functions, and by the fact that the genes belonging to a pathway are scattered over the entire genome. The goal of this thesis was to design, set up and validate a strategy aiming at facilitating the remodelling of (essential) pathways, based on simplifying and reorganizing the yeast genome. The starting point of this research is the central carbon metabolism and in particular, as proof of concept, the glycolytic pathway.