Analysis of protein glycosylation by mass spectrometry |
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Authors: | Morelle Willy Michalski Jean-Claude |
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Institution: | Unité Mixte de Recherche CNRS/USTL 8576, Glycobiologie Structurale et Fonctionnelle, IFR 147, Batiment C9, Université des Sciences et Technologies de Lille 1, Villeneuve d'Ascq Cedex, France. willy.morelle@univ-lille1.fr |
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Abstract: | 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 beta-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). |
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