Dissecting the liver tumour micro-environment: cell- and subcellular-specific characterisation of N-glycosylated basigin

Basigin (CD147), an N-glycosylated transmembrane protein occurring in four potential isoforms, isexpressed on surfaces of immune and cancer cells forming the heterogeneous tumour microenvironment.Basigin display pro- and anti-tumorigenic functions by interacting with multiplebinding partners; interactions that are modulated by glycosylation. However, the molecular featuresof basigin and the N-glycosylation of the cells of the tumour micro-environment remainundescribed. This thesis aimed to characterise the low abundance basigin from primary neutrophilsand HepG2 cancer cells as representative immune and cancer components of the liver tumourmicro-environment. Multiple enrichment and isolation strategies including immunoprecipitation,cell surface protein capture and plasma membrane separation were performed in conjunction withwestern blotting and advanced tandem mass spectrometry. For the first time, human basiginisoform-2 was successfully characterised from both neutrophils and HepG2 by extensive mappingof the polypeptide chain. Cell- and subcellular-specific N-glycomics revealed paucimannose-rich Nglycosylationof neutrophil plasma membranes and dominant high mannose type N-glycans ofHepG2 microsomes. In conclusion, glycobiology and powerful glyco-analytical technologies werebrought together to reveal novel molecular features of a key cancer and immune glycoprotein.These findings contribute to our understanding of the structure and function of basigin andsubcellular-specific N-glycosylation in the tumour micro-environment. (200 words)