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Direct in-gel fluorescence detection and cellular imaging of O-GlcNAc-modified proteins

Clark, Peter M. and Dweck, Jessica F. and Mason, Daniel E. and Hart, Courtenay R. and Buck, Suzanne B. and Peters, Eric C. and Agnew, Brian J. and Hsieh-Wilson, Linda C. (2008) Direct in-gel fluorescence detection and cellular imaging of O-GlcNAc-modified proteins. Journal of the American Chemical Society, 130 (35). pp. 11576-11577. ISSN 0002-7863. PMCID PMC2649877. doi:10.1021/ja8030467.

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We report an advanced chemoenzymatic strategy for the direct fluorescence detection, proteomic analysis, and cellular imaging of O-GlcNAc-modified proteins. O-GlcNAc residues are selectively labeled with fluorescent or biotin tags using an engineered galactosyltransferase enzyme and [3 + 2] azide−alkyne cycloaddition chemistry. We demonstrate that this approach can be used for direct in-gel detection and mass spectrometric identification of O-GlcNAc proteins, identifying 146 novel glycoproteins from the mammalian brain. Furthermore, we show that the method can be exploited to quantify dynamic changes in cellular O-GlcNAc levels and to image O-GlcNAc-glycosylated proteins within cells. As such, this strategy enables studies of O-GlcNAc glycosylation that were previously inaccessible and provides a new tool for uncovering the physiological functions of O-GlcNAc.

Item Type:Article
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URLURL TypeDescription Information CentralArticle
Hsieh-Wilson, Linda C.0000-0001-5661-1714
Additional Information:© 2008 American Chemical Society. Received April 24, 2008. Publication Date (Web): August 7, 2008. We thank Dr. P. Qasba for providing the mutant GalT plasmid, J. Rexach for providing neurons and helpful discussions, T. Nyberg for helpful discussions, and Prof. N. Pierce for use of the FujiFilm imager. This work was supported by the NIH (RO1 GM084724) and Howard Hughes Medical Institute.
Funding AgencyGrant Number
NIHRO1 GM084724-05
Howard Hughes Medical Institute (HHMI)UNSPECIFIED
Issue or Number:35
PubMed Central ID:PMC2649877
Record Number:CaltechAUTHORS:20150519-144346544
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Official Citation:Direct In-Gel Fluorescence Detection and Cellular Imaging of O-GlcNAc-Modified Proteins Peter M. Clark, Jessica F. Dweck, Daniel E. Mason, Courtenay R. Hart, Suzanne B. Buck, Eric C. Peters, Brian J. Agnew, and Linda C. Hsieh-Wilson Journal of the American Chemical Society 2008 130 (35), 11576-11577 DOI: 10.1021/ja8030467
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:57671
Deposited By: Tony Diaz
Deposited On:19 May 2015 23:55
Last Modified:10 Nov 2021 21:53

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