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Analysis of protein glycosylation by mass spectrometry

2007; Nature Portfolio; Volume: 2; Issue: 7 Linguagem: Inglês

10.1038/nprot.2007.227

1754-2189

Willy Morelle, Jean‐Claude Michalski,

RNA and protein synthesis mechanisms

We present a detailed protocol for the structural analysis of protein-linked glycans. In this approach, appropriate for glycomics studies, N-linked glycans are released using peptide N-glycosidase F and O-linked glycans are released by reductive alkaline β-elimination. Using strategies based on mass spectrometry (matrix-assisted laser desorption/ionization–time of flight mass spectrometry and nano-electrospray ionization mass spectrometry/mass spectrometry (nano-ESI-MS-MS)), chemical derivatization, sequential exoglycosidase digestions and linkage analysis, the structures of the N- and/or O-glycans are defined. This approach can be used to study the glycosylation of isolated complex glycoproteins or of numerous glycoproteins encountered in a complex biological medium (cells, tissues and physiological fluids).