The trend away from fossil-fuels toward renewable fuels has boosted the production of biodiesel in the last years. Concurrently, large quantities of glycerol, the main by-product of biodiesel production, are generated. Although pure glycerol finds many applications in the food and pharmaceutical industry, costly and energy-consuming processes are required to purify the crude glycerol beforehand. Therefore, an interesting alternative is the direct bioconversion of crude glycerol to value-added products by microorganisms which can utilize glycerol without the need for expensive refinements.

One fascinating microorganism known to grow well on glycerol is the oleaginous yeast Yarrowia lipolytica. Not only does Y. lipolytica show fast growth rates on glycerol but it is also able to convert it with a high metabolic flexibility into valuable metabolites. Depending on the strain and cultivation condition, Y. lipolytica produces fatty acids, organic acids (e.g. citric acid, ketoglutaric acid), polyols (e.g. arabitol, erythritol, mannitol) and extracellular enzymes (e.g. lipases, proteases). Thus, Y. lipolytica represents an interesting cell factory for the production of industrially relevant products from glycerol.

To obtain economically viable bioprocesses, high product titres, product yields and fast substrate to product conversion rates are required. Meeting these requirements often necessitates a combination of bioprocess engineering and metabolic engineering approaches. For metabolic engineering, it is essential to know the proteins involved in certain metabolic pathways to be able to systematically engineer them to improve the glycerol uptake, conversion and product secretion. Although the understanding of the glycerol metabolism of Y. lipolytica is ever-growing, many enzymes, transcription factors and transport proteins are still unknown.

The aim of this PhD thesis is to identify targets for metabolic engineering of the glycerol metabolism in Y. lipolytica. Therefore, proteins that are putatively involved in the glycerol metabolism will be selected based on similarity to known fungal proteins, on predicted yet not characterized proteins or via differential gene expression analysis. Since the metabolism depends on the environmental conditions, the mutant strains will be cultivated under controlled conditions in bioreactors to evaluate the effect of knocking-out or overexpressing the selected genes on the phenotype of Y. lipolytica. Hence, these results will potentially enable to further conclude on the function of the respective proteins in the glycerol metabolism. Moreover, the influence of certain environmental factors, such as nitrogen availability, aeration or pH, on the metabolism will be analysed to aid the development of a bioprocess and a strain that are adapted to each other.

This PhD project is conducted within the framework of the CD Laboratory for Biotechnology of Glycerol, which focuses on the microbial conversion of raw glycerol into valuable substances.