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Title: Structural analysis of bacterial glycoconjugates
Author: Yang, Tiandi
Awarding Body: Imperial College London
Current Institution: Imperial College London
Date of Award: 2016
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Bacterial glycoconjugates are a group of highly complex macromolecules that are deeply implicated in various biological processes. ey can be crudely divided into several subgroups mainly including bacterial glycolipids and glycoproteins. eir structural complexity once hindered the elucidation of the biological roles they are playing. Based on the development of modern chromatography, mass spectrometry and nuclear magnetic resonance, research described in this thesis analysed the structures of multiple glycoconjugates such as glycolipids and glycoproteins. Lipopolysaccharides are characteristic bacterial glycolipids. e Pseudomonas aeruginosa lipid A and Helicobacter pylori lipopolysaccharide was investigated by a combination of chemical, MS and/or NMR strategies. For the lipid A, it was proved to be modi ed by 4-amino- arabinose, which is associated with bacterial antibiotic resistance. For the lipopolysaccharide, the speci cities of mutiple putative glycosyltransferases, including HP1284, HP1283, HP1578, HP0102 and WaaL ligase, involved in its biosynthesis were characterised. e structural data suggest Helicobacter pylori WaaL ligase has a relaxed speci city in vivo that transfers an un- expected long O-antigen to a short core-oligosaccharide. In addition, our data shed light on the regulation of bacterial O-antigen biosynthesis. e heavily glycosylated mbrial-associated protein 1 of Streptococcus parasanguinis was subjected to in-depth glycomics and glycoproteomics, which helps determine the speci cities of multiple putative glycosyltransferases such as GALT1/2 and GLY. More speci cally, GALT1 was proved to be a bi-functional glycosyltransferase that transfers a glucose and an N-acetyl- glucosamine. GALT2 and GLY was deteremined to transfer a glucose and a rhamnose, respectively. Based on these conclusions, our study helps elucidate the glycosylation pathway of mbrial-associated protein 1. e recent characterisation of bacterial oligosaccharyltransferases has the potential to revolutionise the production of glycoconjugate vaccines. e putative O-linked oligosaccharyltransferases system in Burkholderia thailandensis was investigated based on a zwitterionic hydrophilic interaction liquid chromatography mass spectrometry method, which helps expand the toolbox of glyco-engineering. By using a simpilied liquid chromatography- mass spectrometry method, multiple glycoproteins in Coxiella burnetii were semi-identi ed. During these studies, many classical chemistries were resurrected to inspire novel chemical strategies and facilitate the instrumental analysis. We hope the methods included in this thesis can contribute to further research where comprehensive structural characterisations of complex bacterial glycoconjugates are required.
Supervisor: Dell, Anne Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral