Analysis of protein glycosylation by mass spectrometry.
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).
Morelle
Willy
W
Unité Mixte de Recherche CNRS/USTL 8576, Glycobiologie Structurale et Fonctionnelle, IFR 147, Bâtiment C9, Université des Sciences et Technologies de Lille 1, Villeneuve d'Ascq Cedex, France. willy.morelle@univ-lille1.fr
Michalski
Jean-Claude
JC
D006023:Glycoproteins / Q000737:chemistry*; D006031:Glycosylation; D013058:Mass Spectrometry / Q000379:methods*; D015394:Molecular Structure; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase / Q000378:metabolism; D011134:Polysaccharides / Q000032:analysis* / Q000737:chemistry*
D006023:Glycoproteins; D011134:Polysaccharides; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase