Background The oleaginous yeast Cutaneotrichosporon oleaginosus represents one of the most promising microbial platforms for resource-efficient and scalable lipid production, with the capacity to accept a wide range of carbohydrates encapsulated in complex biomass waste or lignocellulosic hydrolysates. Currently, data related to molecular aspects of the metabolic utilisation of oligomeric carbohydrates are sparse. In addition, comprehensive proteomic information for C. oleaginosus focusing on carbohydrate metabolism is not available. Results In this study, we conducted a systematic analysis of carbohydrate intake and utilisation by C. oleaginosus and investigated the influence of different di- and trisaccharide as carbon sources. Changes in the cellular growth and morphology could be observed, depending on the selected carbon source. The greatest changes in morphology were observed in media containing trehalose. A comprehensive proteomic analysis of secreted, cell wall-associated, and cytoplasmatic proteins was performed, which highlighted differences in the composition and quantity of secreted proteins, when grown on different disaccharides. Based on the proteomic data, we performed a relative quantitative analysis of the identified proteins (using glucose as the reference carbon source) and observed carbohydrate-specific protein distributions. When using cellobiose or lactose as the carbon source, we detected three- and five-fold higher diversity in terms of the respective hydrolases released. Furthermore, the analysis of the secreted enzymes enabled identification of the motif with the consensus sequence LALL[LA]L[LA][LA]AAAAAAA as a potential signal peptide. Conclusions Relative quantification of spectral intensities from crude proteomic datasets enabled the identification of new enzymes and provided new insights into protein secretion, as well as the molecular mechanisms of carbo-hydrolases involved in the cleavage of the selected carbon oligomers. These insights can help unlock new substrate sources for C. oleaginosus, such as low-cost by-products containing difficult to utilize carbohydrates. In addition, information regarding the carbo-hydrolytic potential of C. oleaginosus facilitates a more precise engineering approach when using targeted genetic approaches. This information could be used to find new and more cost-effective carbon sources for microbial lipid production by the oleaginous yeast C. oleaginosus.

The oleaginous yeast Yarrowia lipolytica is capable of accumulating large amounts of fatty acids in the form of lipids which can serve as a platform polymer for a variety of applications. Additionally, through heterologous gene expression, Y. lipolytica is capable of producing carotenoids. Due to the observation that Y. lipolytica exhibits a high phenotypic inter-strain variability, robotics and microwell scale cultivations are applied to examine 12 wild type strains of a Y. lipolytica collection. The strains are characterized with respect to their metabolic capabilities for fatty acid production as well as genetically manipulated to produce beta-carotene. Furthermore, growth and production behavior of the strain collection at different temperatures as well as initial pH are assessed. A molecular discrimination between the strains is achieved by internal transcribed spacer (ITS)-sequencing and polymerase chain reaction (PCR)-based methods. The best performing strain with respect to lipid production produces ≈2% lipids per dry cell weight (DCW) and 8 mg g −1 beta-carotenoids in yeast nitrogen base (YNB) media. All strains show robust growth from 28 to 34°C. Practical Applications: Unlocking nature's potential by screening natural isolates shows that even on inter-strain level sufficient phenotypic diversity may arise. Automated growth-based characterization of beta-caroteneproducing strains in terms of media composition, effect of initial pH, and temperature tolerance shows that with modern cultivation techniques, rapid characterization of strain collections is feasible. Combining results of beta-carotene and lipid formation could help to balance both pathways to improve the storage of hydrophobic compounds in the lipid droplets. The generalized findings could be further transferred to improve the production of any valuable compound, derived from the mevalonate pathway.

Plant based triacylglycerides are a sustainable feedstock for the chemical, bioenergy, food and cosmetic sector. While fatty acids conversion has been intensively studied, processes for glycerol valorization have been scarce. In contrast to chemical hydrolysis of plant triacylglycerides enzymatic hydrolysis methods provide a cleaner glycerol stream. This study focuses on the selection of a commercial lipase capable of quantitative hydrolysis of rapeseed‐ and high oleic sunflower oils. Enzymatic process using only water as the reaction medium allows simplified recovery of pure glycerol. Among the six commercial lipase preparations tested, Candida rugosa lipase was identified as the most effective biocatalyst. Catalytic behavior in buffer and pure water was equivalent. Glycerol generated using a washed lipase was recovered by just three purification steps. The FTIR spectrum of the recovered glycerol was equivalent to pure glycerol standard. Over the entire recovery process, 82%–88% of the theoretical glycerol yield could be obtained. Purified glycerol was further didehydroxylated to allyl alcohol by a formic acid mediated open distillation process. The enriched allyl alcohol had a quality, which allows to use it for the synthesis of bio‐based acrylonitrile.