Purification and carbohydrate structure of natural murine interferon-beta.
Mouse interferon-beta (Mu-INF-beta) induced in C-243 cells with Newcastle disease virus was purified in four steps including ammonium sulfate fractionation. DEAE-cellulose, monoclonal Mu-IFN-beta antibody affinity and Mono-S cation-exchange chromatographies. Specific activity of the purified Mu-IFN-beta ranged over 1.1-1.4 X 10(9) NIH units/mg protein. This preparation was submitted to pronase digestion and gel 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-methylgalactose; 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-INF-beta: (formula; see text).
Civas
A
A
Unité Associée 040622 du Centre National de la Recherche Scientifique, Université de Paris XI, Chatenay-Malabry, France.
Fournet
B
B
Coulombel
C
C
Le Roscouet
D
D
Honvault
A
A
Petek
F
F
Montreuil
J
J
Doly
J
J
D000818:Animals; D002240:Carbohydrate Sequence; D002241:Carbohydrates / Q000302:isolation & purification*; D002850:Chromatography, Gel; D006863:Hydrogen-Ion Concentration; D007370:Interferon Type I / Q000302:isolation & purification*; D013058:Mass Spectrometry; D008745:Methylation; D051379:Mice; D008969:Molecular Sequence Data
D002241:Carbohydrates; D007370:Interferon Type I