Maximilian Aßkamp

• The valorization of carbon-rich industrial waste-streams, such as crude glycerol derived from the biodiesel industry, is a promising avenue to overcome the dependence on fossil resources for the production of certain chemicals. Glycerol has a higher degree of reduction compared to sugars, allowing higher theoretical yields of small reduced molecules, when used as a substrate in industrial biotechnological processes. Several microorganisms are able to catabolize glycerol, including the Saccharomyces cerevisiae strain CBS 6412-13A. In order to utilize glycerol’s degree of reduction for S. cerevisiae-based synthesis of fermentation products, metabolic engineering is required. A replacement of the native FAD-dependent L-glycerol 3-phosphate pathway by the artificial NADH-delivering dihydroxyacetone pathway is supposed to provide glycerol-derived electrons for the cytosolic formation of fermentation products. However, S. cerevisiae has potent respiratory mechanisms, that maintain cytosolic redox balance by transferring electrons from cytosolic NADH to oxygen. During growth with e.g. ethanol or low glucose concentrations, the external NADH dehydrogenases (Nde1/2) and the L-G3P shuttle (composed of Gpd1/2 and Gut2), are contributing to cytosolic NAD+ regeneration. Their contribution with glycerol as carbon source was not investigated so far. It is demonstrated here, that in particular Nde1 significantly contributes to cytosolic redox balance in S. cerevisiae catabolizing glycerol. S. cerevisiae’s major native fermentative mechanism to oxidize NADH is alcoholic fermentation, even when oxygen is available. However, glycerol is generally considered to be a ‘non-fermentable’ carbon source for S. cerevisiae. The current work demonstrates that at least S. cerevisiae strains catabolizing glycerol via the DHA pathway are able to perform alcoholic fermentation. The results of this thesis provide important insights into the redox metabolism of S. cerevisiae catabolizing glycerol.