Characterization ofO-Glycosylated Precursors of Insulin-like Growth Factor II by Matrix-assisted Laser Desorption/Ionization Mass Spectrometry

✍ Scribed by Jespersen, S.; Koedam, J. A.; Hoogerbrugge, C. M.; Tjaden, U. R.; van der Greef, J.; Van den Brande, J. L.

• Publisher: John Wiley and Sons
• Year: 1996
• Tongue: English
• Weight: 748 KB
• Volume: 31
• Category: Article
• ISSN: 1076-5174
• DOI: 10.1002/(sici)1096-9888(199608)31:8<893::aid-jms374>3.0.co;2-s

✦ Synopsis

High molecular weight precursors of insulin-like growth factor 11 (IGF-11) were isolated from Cohn fraction IV of human plasma by uitrafiltration, affinity chromatography and reversed-phase high-performance liquid chromatography. Molecular weight determination by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) of two high molecular weight IGF-I1 preparations revealed heterogeneous glycosylation. A combination of enzymatic degradation and MALDI-MS were applied for further structural characterization of the glycosylated precursors of IGF-11. The first step was molecular weight determination of intact high molecular weight IGF-11s prior to and after treatment with neuraminidase and 0-glycosidase. This, together with a comparison of molecular weight information available from the cDNA, revealed that both high molecular weight IGF-I1 species contain an identical C-terminal extension of 20 residues but different degrees of glycosylation. Second, comparative Eodo Glu-C digestion of the preparations prior to and after enzymatic release of carbohydrates and subsequent remeasurement of the molecular weight by MALDI-MS confirmed the primary structure of precursor IGF-II'-*'. The 0-linked carbohydrates were found to be associated with the C-terminal extension and the heterogeneity was identified as varied sialylated forms of one and two HexNAc-Hex groups.