Characterization of O-glycosylated precursors of insulin-like growth factor II by matrix-assisted laser desorption/ionization mass spectrometry
article
High molecular weight precursors of insulin-like growth factor II (IGF-II) were isolated from Cohn fraction IV of human plasma by ultrafiltration, 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-II preparations revealed heterogeneous glycosylation. A combination of enzymatic degradation and MALDI-MS were applied for further structural characterization of the glycosylated precursors of IGF-II. The first step was molecular weight determination of intact high molecular weight IGF-IIs prior to and after treatment with neuraminidase and O-glycosidase. This, together with a comparison of molecular weight information available from the cDNA, revealed that both high molecular weight IGF-II species contain an identical C-terminal extension of 20 residues but different degrees of glycosylation. Second, comparative Endo 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-II1-87. The O-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.
Topics
Matrix-assisted laser desorption/ionizationO-glycosylationPrecursor insulin-like growth factor IISomatomedin bAmino acid sequenceArticleDrug isolationDrug purificationDrug structureGlycosylationMass spectrometryMolecular weightPriority journalProtein degradationAmino Acid SequenceGlycoside HydrolasesHumansHydrolysisInsulin-Like Growth Factor IIMolecular Sequence DataMolecular WeightNeuraminidaseOligosaccharidesPeptide MappingProtein PrecursorsSpectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
TNO Identifier
233414
ISSN
10765174
Source
Journal of Mass Spectrometry, 31(8), pp. 893-900.
Pages
893-900
Files
To receive the publication files, please send an e-mail request to TNO Repository.