Carbohydrates are the most diverse family of biomolecules in nature. From a panel of mono-saccharides organisms build a vast variety of glycans and glycoconjugates with essential biological functions in energy metabolism, cellular communication and structural integrity to name a few. The wide array of architectures found in glycans is orchestrated by carbohydrate active enzymes which control glycosidic bonds between mono-saccharides and perform additional carbohydrate modifications. In order to assess biological functions, structural information is essential as is the ability to modify carbohydrates to use as biological probes. This thesis addresses analytical tools to gain insights into carbohydrates and the enzymes involved in the glycan metabolism. An easy NMR assay is presented to monitor enzymatic oligo-saccharide oxidation with minimal sample preparation, while the regio-selective oxidation products are valuable targets as well as precursors for industrial biotechnology applications. The adaptation of a glyco array platform for rapid screening for glycoside hydrolase activities of fungal enzymes towards mixed oligo-saccharide libraries advances the analytical possibilities and provides a tool for the identification of novel enzymatic activities. While state-of-the-art N-glycan analysis solves the problem of isobaric linkage isomers through the application of ion mobility, traditional methods heavily rely on exo-glycoside hydrolases. The discovery and proven applicability of an Î±2,6-'pseudosialidase' completes the analytical toolbox for N-acetylneuraminic acid terminated N-glycans. The identification of glucosylsphingosine as an endogenous modulator of DAT-mediated dopamine transport is an exciting discovery and may reveal a new dimension to the etiology of Parkinson's disease. The methods presented in this thesis provide glycoscientists with tools to further analyse glycans, CAZymes and their impact on biotechnology.
|Date of Award||1 Aug 2018|
- The University of Manchester
|Supervisor||Sabine Flitsch (Supervisor) & Perdita Barran (Supervisor)|
- Parkinson's disease
- galactose oxidase