Glycosylation of extracellular superoxide dismutase studied by high-performance liquid chromatography and mass spectrometry.
Extracellular superoxide dismutase, EC-SOD, the main superoxide dismutase in biological fluids, is known from its lectin binding to be a glycoprotein. We have characterized the glycosylation of recombinant EC-SOD. A tryptic digest of the protein contained only one glycosylated peptide. This peptide was specifically bound to lectins and stained by periodic acid-Schiff stain. Although appearing very large on size-exclusion chromatography, it was shown to be glycosylated at only one site, asparagine-89, by specific cleavage with glycanases followed by mass spectrometry of the resulting peptide. Based on the binding properties of the peptide to concanavalin A and lentil lectin and the elution profile of N-glycanase-treated glycopeptide on ion-exchange chromatography, the carbohydrate appears to be the complex biantennary type with a core fucose.
Strömqvist
M
M
Symbicom AB, Umeå, Sweden.
Holgersson
J
J
Samuelsson
B
B
D000596:Amino Acids / Q000032:analysis; D002240:Carbohydrate Sequence; D002241:Carbohydrates / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D006020:Glycopeptides / Q000302:isolation & purification; D006868:Hydrolysis; D037102:Lectins; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010503:Periodic Acid-Schiff Reaction; D011994:Recombinant Proteins / Q000302:isolation & purification; D013482:Superoxide Dismutase / Q000737:chemistry* / Q000302:isolation & purification; D014357:Trypsin
D000596:Amino Acids; D002241:Carbohydrates; D006020:Glycopeptides; D037102:Lectins; D011994:Recombinant Proteins; D013482:Superoxide Dismutase; D014357:Trypsin