Incorporation of Unnatural Sugars for the Identification of Glycoproteins

  • Balyn W. Zaro
  • Howard C. Hang
  • Matthew R. Pratt
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 951)

Abstract

Glycosylation is an abundant post-translational modification that alters the fate and function of its substrate proteins. To aid in understanding the significance of protein glycosylation, identification of target proteins is key. As with all proteomics experiments, mass spectrometry has been established as the desired method for substrate identification. However, these approaches require selective enrichment and purification of modified proteins. Chemical reporters in combination with bioorthogonal reactions have emerged as robust tools for identifying post-translational modifications including glycosylation. We provide here a method for the use of bioorthogonal chemical reporters for isolation and identification of glycosylated proteins. More specifically, this protocol is a representative procedure from our own work using an alkyne-bearing O-GlcNAc chemical reporter (GlcNAlk) and a chemically cleavable azido-azo-biotin probe for the identification of O-GlcNAc-modified proteins.

Key words

Proteomics Glycosylation Bioorthogonal chemical reporter Click chemistry Azide Mass spectrometry O-GlcNAc 

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Copyright information

© Springer Science+Business Media, LLC 2013

Authors and Affiliations

  • Balyn W. Zaro
    • 1
  • Howard C. Hang
    • 2
  • Matthew R. Pratt
    • 1
    • 3
  1. 1.Department of ChemistryUniversity of Southern CaliforniaLos AngelesUSA
  2. 2.The Laboratory of Chemical Biology and Microbial PathogenesisThe Rockefeller UniversityNew YorkUSA
  3. 3.Department of Molecular and Computational BiologyUniversity of Southern CaliforniaLos AngelesUSA

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