Typescript. ; "Submission date: 13th May 2010" ; Thesis by publication. ; Includes one CD-Rom in back pocket. ; CD-Rom contains supplementary tables in .doc, .pdf and .xls formats. ; Thesis (PhD)--Macquarie University, Faculty of Science, Dept. of Chemistry and Biomolecular Sciences, 2010. ; Bibliography: pages 293-313. ; 1. Introduction -- 2. General methods -- 3. Approaches for membrane glycoprotein analysis -- 4. Liver membrane proteome glycosylation changes in mice bearing an extra-hepatic tumour -- 5. Effects of a distal colon 26 adenocarcinoma on the liver membrane proteome of non-cachectic and cachectic mice -- 6. General discussion -- Appendices ; The study of membrane proteins has become increasingly important, as they play a number of cellular roles such as signal transduction, cell adhesion and cell communication. However, membrane proteomics has often been hampered by the low abundance, extreme hydrophobicity and large size of membrane proteins and therefore alternative enrichment strategies are required to efficiently analyse these types of proteins. In this thesis, we developed an alternative strategy for enriching membrane proteins by targeting both the hydrophobicity and glycosylation of these proteins, which involved (1) Triton X-114 detergent phase partitioning (2) isolation of glycosylphosphatidylinositol (GPI)-anchored proteins and (3) glycoprotein capture using either lectin affinity or hydrazine chemistry. Using this strategy we identified four predicted GPI-anchored proteins and two glycoproteins (Limbic system-associated membrane protein precursor (LSAMP) and Neuronal growth regulator 1 precursor) that have not previously been seen in the liver. ; The capture of membrane glycoproteins by lectin affinity and hydrazine chemistry resulted in mostly different populations of enriched glycoproteins demonstrating the complementarity of the two glycocapture approaches. Lectins are often preferred as tools for studying glycoproteins because the sugars on the enriched proteins remain intact compared to the hydrazine capture approach. The selectivity and specificity of lectins for specific glycan structures however, was not reflected in the glycoforms retained from a complex mixture of proteins such as those present in the liver membrane. The use of ConA and WGA for enrichment of membrane glycoproteins showed little difference between the N-glycans on the bound proteins compared to those unbound. Only Jacalin appeared to enrich for O-glycan containing glycoproteins from the liver membrane preparation. Lectins are valuable tools for enrichment and fractionation of glycoproteins; however detailed structural analysis must be undertaken to define specific differences in the glycosylation of the proteins of interest in a complex mixture. ; Applying label-free shotgun proteomics we determined that the presence of a distal Engelbreth-Holm Swarm (EHS) sarcoma and a distal colon 26 (C26) adenocarcinoma could induce differential expression of many metabolic classes of hepatic enzymes of cancer bearing mice, demonstrating the extensive systemic effect of a distal cancer on the functions of the liver. In particular, we determined that many enzymes involved in the protein glycosylation machinery in the liver were differentially expressed in cancer-bearing mice. Glycomics analysis determined that differential expression of these enzymes was reflected in changes to specific glycan structures expressed on the liver membrane glycoproteins. Both distal cancer models (EHS sarcoma and C26 adenocarcinoma) resulted in an overall increased sialylation of N-glycans presented on the liver cell membrane proteins compared to the control. The liver membrane glycoproteins of EHS-bearing mice showed increased sialylation of core 1 and core 2 O-glycans containing NeuGc residues, whilst there were no observable O-glycan differences observed on the liver membrane proteins of non-cachectic and cachectic C26-bearing mice. Specifically, both EHS bearing mice and non-cachectic mice bearing the C26 adenocarcinoma were shown to preferentially display more N-acetyl neuraminic acids (NeuAc) than N-glycolylneuraminic acid (NeuGc) on their N-glycans. The preferred expression of NeuAc-containing glycans over the more common NeuGc-containing glycans suggests that the CMP-hydroxylase enzyme, which converts NeuAc to NeuGc, had impaired function and/or expression in these mice. There was also increased expression of N-glycan structures containing NeuAcα(2-3/6)Galβ(1-4)GlcNAcβ(1-2) (sialyl LacNAc) on the liver proteins in both cancer-bearing murine models compared to control livers. The expression of this specific glycan motif on the liver cell membrane of mice bearing two different distal cancers suggests that this glycan motif may serve as a marker for tumour-induced liver inflammation in the mouse. ; The differential expression of the enzymes in the glycosylation pathway and the corresponding glycan products observed on the liver membrane glycoproteins of EHS-bearing and C26-bearing mice strongly suggests that glycosylation of liver membrane proteins plays a role in the systemic inflammatory responses in the liver to the presence of distal cancers. ; Mode of access: World wide web ; 1 online resource (314 pages) illustrations (some colour)