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A Boronic Acid-Based Enrichment for Site-Specific Identification of the N-glycoproteome Using MS-Based Proteomics

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Analysis of Post-Translational Modifications and Proteolysis in Neuroscience

Part of the book series: Neuromethods ((NM,volume 114))

Abstract

Modification of proteins by N-linked glycans plays a critically important role in biological systems, including determining protein folding and trafficking as well as regulating many biological processes. Aberrant glycosylation is well known to be related to disease, including cancer and neurodegenerative diseases. Current mass spectrometry (MS)-based proteomics provides the possibility for site-specific identification of the N-glycoproteome; however, this is extraordinarily challenging because of the low abundance of many N-glycoproteins and the heterogeneity of glycans. Effective enrichment is essential to comprehensively analyze N-glycoproteins in complex biological samples. The covalent interaction between boronic acid and cis-diols allows us to selectively capture glycopeptides and glycoproteins, whereas the reversible nature of the bond enables them to be released after non-glycopeptides are removed. By virtue of the universal boronic acid-diol recognition, large-scale mapping of N-glycoproteins can be achieved by combining boronic acid-based enrichment, PNGase F treatment in the presence of heavy oxygen (18O) water, and MS analysis. This method can be extensively applied for the comprehensive analysis of N-glycoproteins in a wide variety of complex biological samples.

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Acknowledgements

This work is supported by the National Science Foundation (CAREER Award, CMI-1454501).

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Authors and Affiliations

  1. School of Chemistry and Biochemistry, Georgia Institute of Technology, Atlanta, GA, 30332, USA

    Haopeng Xiao, George X. Tang, Weixuan Chen & Ronghu Wu

  2. Petit Institute for Bioengineering and Bioscience, Georgia Institute of Technology, Atlanta, GA, 30332, USA

    Haopeng Xiao, George X. Tang, Weixuan Chen & Ronghu Wu

Authors
  1. Haopeng Xiao
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  2. George X. Tang
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  3. Weixuan Chen
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  4. Ronghu Wu
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Corresponding author

Correspondence to Ronghu Wu .

Editor information

Editors and Affiliations

  1. Department of Biology, University of Wisconsin-Stout, Menomonie, Wisconsin, USA

    Jennifer Elizabeth Grant

  2. Department of Microbiology, Biochemistry and Molecular Genetics, Rutgers University—New Jersey Medical School Cancer Center Newark, Newark, New Jersey, USA

    Hong Li

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Xiao, H., Tang, G.X., Chen, W., Wu, R. (2015). A Boronic Acid-Based Enrichment for Site-Specific Identification of the N-glycoproteome Using MS-Based Proteomics. In: Grant, J., Li, H. (eds) Analysis of Post-Translational Modifications and Proteolysis in Neuroscience. Neuromethods, vol 114. Humana Press, New York, NY. https://doi.org/10.1007/7657_2015_94

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  • DOI: https://doi.org/10.1007/7657_2015_94

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3470-6

  • Online ISBN: 978-1-4939-3472-0

  • eBook Packages: Springer Protocols

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