Summary
Secreted as well as membrane-associated eukaryotic proteins are most commonly glycosylated. Saccharides are attached to proteins mainly through N-and O-glycosydic bonds or as part of the glycosylphosphatidyinositol-membrane anchor. In contrast to N-glycosylation, which involves the co-translational transfer in the endoplasmic reticulum (ER) of the glycan portion of Glc3Man9GlcNAc2-PP-dolichol to suitable Asn residues on nascent polypeptides, O-glycosylation begins with the addition of a single monosaccharide. Contrary to N-glycosylation, which involves an asparagine residue in the sequon Asn-Xaa-Thr/Ser (Xaa can be any amino acid except Pro, and it is rarely Cys), no particular sequence motif has been described for O-glycosylation. This may reflect the fact that: (1) the specificity of the UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferase is presently unknown; and (2) seems to be modulated by sequence context, secondary structure, and surface accessibility (1). An internet server, accessible at http://www.cbs.dtu.dk/netOglyc/cbsnetOglyc.html, produces neural network predictions of mucin-type GalNAc O-glycosylation sites in mammalian proteins based on 299 known and verified mucin-type O-glycosylation sites. The sequence context of glycosylated threonines was found to differ from that of serine, and the sites were found to cluster. Nonclustered sites had a sequence context different from that of clustered sites, and charged residues were disfavored at position −1 and +3.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Hansen, J. E., Lund, O., Tolstrup, N., Gooley, A. A., Williams, K. L., and Brunak, S. (1998) NetOglyc: Prediction of mucin type O-glycosylation sites based on sequence context and sur face accessibility. Glycoconjugate J., 15, 115–130.
van den Steen, P., Rudd, P. M., Dwek, R. A., and Opdenakker, G. (1998) Concepts and principles of O-linked glycosylation. Crit. Rev. Biochem. Mol. Biol. 33, 151–208.
Gemmill, T. R. and Trimble, R. B. (1999) Overview of N- and O-linked oligosaccharide struc tures in various yeast species. Biochim. Biophys. Acta 1426, 227–237.
Strahl-Bolsinger, S., Gentzsch, M., and Tanner, W. (1999) Protein O-mannosylation. Biochim. Biophys. Acta 1426, 297–307.
Hounsell, E. F., Davies, M. J., and Renouf, D. V. (1996) O-linked protein glycosylation struc ture and function. Glycoconjugate J., 13, 19–26.
Fountoulakis, M. and Lahm, H-W. (1998) Hydrolysis and amino acid composition of pro teins. J. Chromatogr. A, 826, 109–134.
Whittal, R. M., Palcic, M. M., Hindsgaul, O., and Li, L. (1995) Direct analysis of enzymatic reactions of oligosaccharides in human serum using matrix-assisted laser desorption ioniza tion mass spectrometry. Anal. Chem., 67, 3509–3514.
Eirín, M. T., Calvete, J. J., and González-odríguez, J. (1986) New isolation procedure and fur ther biochemical characterization of glycoproteins IIb and IIIa from human platelet plasma membrane. Biochem. J., 240, 147–153.
Calvete, J. J. and González-Rodríguez, J. (1986) Isolation and biochemical characterization of the α-and β-subunits of glycoprotein IIb of human platelet plasma membrane. Biochem. J. 240, 155–161.
Montreuil, J., Bouquelet, S., Debray, H., Lemoine, J., Michalski, J-C, Spik, G, and Strecker, G. (1994) Glycoproteins in Carbohydrate Analysis. A Practical Approach (Chaplin, M. F. and Kennedy, J. F., eds.) IRL Press, Oxford, pp. 181–293.
Dell, A., Khoo, K-H, Panico, M., McDowell, R. A., Etienne, A. T, Reason, A. J., and Morris, H. R. (1993) FAB-MS and ES-MS of glycoproteins in Glycobiology. A Practical Approach (Fukuda, M. and Kobata, A., eds.) IRL Press, Oxford, pp. 187–222.
Alving, K., Paulsen, H, and Peter-Katalinic, J. (1999) Characterization of O-glycosylation sites in MUC2 glycopeptides by nanoelectrospray QTOF mass spectrometry. J. Mass Spectrom., 34, 395–407.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2008 Humana Press, a part of Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Calvete, J.J., Sanz, L. (2008). Analysis of O-Glycosylation. In: Kannicht, C. (eds) Post-translational Modifications of Proteins. Methods in Molecular Biology™, vol 446. Humana Press. https://doi.org/10.1007/978-1-60327-084-7_20
Download citation
DOI: https://doi.org/10.1007/978-1-60327-084-7_20
Publisher Name: Humana Press
Print ISBN: 978-1-58829-719-8
Online ISBN: 978-1-60327-084-7
eBook Packages: Springer Protocols