Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.684805
Title: N-linked protein glycosylation in Campylobacter and Helicobacter species
Author: Frost, Helen
ISNI:       0000 0004 5922 7859
Awarding Body: University of Manchester
Current Institution: University of Manchester
Date of Award: 2016
Availability of Full Text:
Access through EThOS:
Access through Institution:
Abstract:
N-linked protein glycosylation is the enzymatic transfer of a carbohydrate glycan to an asparagine residue of a polypeptide, catalysed by an N-oligosaccharyl transferase(OTase). Bacterial N-glycosylation is best understood in the foodborne pathogen Campylobacter jejuni, in which a heptasaccharide glycan is built at cytoplasmic face of the inner membrane, flipped to the periplasm and transferred to a polypeptide enbloc. C. jejuni encodes each of the proteins required for the N-glycosylation pathway in a single genetic region, termed the pgl locus. Homologues of the gene encoding the C. jejuni OTase, PglB, are found in all Campylobacter species, three Helicobacter species, and more distantly related ε- and δ-Proteobacteria species such as Wolinella succinogenes, Desulfovibrio desulfuricans and Nitratiruptor tergarcus. A small numberof Campylobacter species and all three pglB-containing Helicobacter species have two distinct pglB genes, pglB1 and pglB2, along with homologues of other C. jejuni pgl genes. The work presented in this thesis investigated the N-glycosylation system of a bacterial species encoding two distinct PglBs, C. concisus. The roles of the two PglB enzymes in C. concisus were investigated using an in vitro OTase assay, and the structure of a C. concisus N-glycan elucidated by mass spectrometry. The work in this thesis also expands our knowledge of C. jejuni N-glycosylation by investigating the full scope of N-glycosylation using an in silico method to predict the total C. jejuni N-glycoproteome. This was followed by experimental validation of these predictions, in which three novel C. jejuni N-glycoproteins were identified, bringing the number of reported C. jejuni NCTC 11168 N-glycoproteins to 57. One of these novel N-glycoproteins, Cj0633, is the most extensively N-glycosylated bacterial glycoprotein reported to date, with the addition of up to eight N-glycans when expressed in the presence of the C. jejuni pgl machinery. In the final investigation presented in this thesis, a method to identify bacterial N-glycoproteins using anti-glycan antisera to immunoprecipitate N-glycoproteins was developed. These data expand our knowledge of C. concisus N-glycosylation and provide valuable insight into the full scope of N-glycosylation in C. jejuni.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.684805  DOI: Not available
Share: