N-Glycosylation of Recominant Human Interferon-Gamma Produced in Deferent Animal Expression System

Recombinant human interferon-gamma (IFN-gamma) was expressed in Chinese hamster ovary cells, baculovirus-infected Sf9 insect cells and the mammary gland of transgenic mice, The N-linked carbohydrate populations associated with both Asn(25), and Asn(97) glycosylation sites were characterized by matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS) in combination with exoglycosidase array sequencing, a site-specific analysis of dual (2N) and single (1N) site-occupancy variants of IFN-gamma derived from Chinese hamster ovary cells showed that N-glycans were predominantly of the complex bi- and triantennary type, Although Asn(25)-linked glycans were substituted with a core fucose residue, Asn(97) N-glycans were predominantly non-fucosylated, and truncated complex and high-mannose oligosaccharide chains were also evident, Transgenic mouse derived IFN-gamma exhibited considerable site-specific variation in N-glycan structures, Asn(25)-linked carbohydrates were of the complex, core fucosylated type, Asn(97)-linked carbohydrates were mainly of the oligomannose type, with smaller proportions of hybrid and complex N-glycans, Carbohydrates associated with both glycosylation sites of IFN-gamma from Sf9 insect cells were mainly tri-mannosyl core structures, with fucosylation confined to the Asn(25) site, These data demonstrate the profound influence of host cell type and protein structure on the N-glycosylation of recombinant proteins.