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Signal Transduction Protocols pp 175–194Cite as

Nucleocytoplasmic Glycosylation, O-GlcNAc

Identification and Site Mapping

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 284))

Abstract

β-O-linked N-acetylglucosamine (O-GlcNAc) is posttranslationally added to serine and threonine residues of many nuclear and cytoplasmic proteins found in metazoans. This modification is dynamic and responsive to numerous stimuli and conditions, suggesting an important role in many regulatory pathways. Moreover, the O-GlcNAc modification seems to compete with phosphorylation for sites of attachment, indicating a reciprocal relationship with phosphorylation. This chapter includes protocols for: (1) identifying the O-GlcNAc modification on proteins through immunoblotting, lectin affinity chromatography, and galactosyltransferase labeling; and (2) identifying and enriching for the sites of attachment using the mass spectrometry-based β-elimination followed by Michael addition with dithiothreitol (BEMAD) technique.

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Author information

Authors and Affiliations

  1. Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, MD

    Natasha Elizabeth Zachara, Win Den Cheung & Gerald Warren Hart

Authors
  1. Natasha Elizabeth Zachara
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  2. Win Den Cheung
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  3. Gerald Warren Hart
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Editor information

Editors and Affiliations

  1. Department of Molecular and Cellular Biochemistry, College of Medicine, University of Kentucky, Lexington, KY

    Robert C. Dickson  & Michael D. Mendenhall  & 

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© 2004 Humana Press Inc., Totowa, NJ

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Zachara, N.E., Cheung, W.D., Hart, G.W. (2004). Nucleocytoplasmic Glycosylation, O-GlcNAc. In: Dickson, R.C., Mendenhall, M.D. (eds) Signal Transduction Protocols. Methods in Molecular Biology, vol 284. Humana Press. https://doi.org/10.1385/1-59259-816-1:175

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  • DOI: https://doi.org/10.1385/1-59259-816-1:175

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-245-2

  • Online ISBN: 978-1-59259-816-8

  • eBook Packages: Springer Protocols

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