Purification and carbohydrate structure of natural murine interferon-β

Mouse interferon-β (Mu-IFN-β) induced in C-243 cells with Newcastle disease virus was purified in four steps including ammonium sulfate fractionation, DEAE-cellulose, monoclonal Mu-IFN-β antibody affinity and Mono-S cation-exchange chromatographies. Specific activity of the purified Mu-IFN-β ranged over 1.1–1.4 × 109 NIH units/mg protein. This preparation was submitted to pronase digestion and gel filtration on Fractogel TSK HW-40. The permethylated and acetylated glycopeptide fraction was analyzed by chemical-ionization (ammonia) mass spectrometry. The major glycopeptide is composed of Gal, Man, GlcNAc and NeuAc with a molar ratio of 2.0:3.6:3.4:0.5. The GLC pattern of methyl derivatives obtained by methanolysis and acetylation of fully methylated glycopeptide identified 2,3,4,6-tetra-O-methylgalactose; 3,4,6-tri-O-methyl-mannose; 2,3,4-and 2,4,6-tri-O-methyl galactose; 2,4,di-O-methyl mannose and 3,6-di-O-methylglucosamine. These results when compared with data on N-glycans suggest the following structure for the carbohydrate moiety of Mu-IFN-β: