Human interferon a2 (IFN-a2) was expressed in Spodopteru frugiperdu Sf9 insect cells using the baculovirus expression system. The protein purified by immunoaffinity chromatography exhibited biological activity identical to that of leukocyte-derived 'natural' IFN-a2. However, the protein was found to be heterogeneously glycosylated, partially truncated by proteolysis and partially lacking a disulfide bridge. The major product was shown to be 0-glycosylated at the same position as natural human IFN-a2. Enzymatic cleavage, reverse-phase HPLC peptide mapping and plasma-desorption mass spectroscopy analysis revealed the presence of two types of 0-linked carbohydrates. The major 0-linked carbohydrate was found to be the disaccharide galactosyl(/31-3)-N-acetylgalactosamine, the minor component the monosaccharide N-acetylgalactosamine. No evidence for sialylation was found. The non-glycosylated species representing about 40% of the total material were shown to partially lack the C-terminal three amino acids. In addition an unglycosylated, reduction-sensitive dimer was observed. This was formed due to the lack of the N-terminal cysteine normally forming an intramolecular disulfide bridge. Furthermore, a minor species was identified which contains Cysl and Cys98 in a modified form, thereby hindering the formation of a disulfide bridge between these two residues.
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