Identification of new O-GlcNAc modified proteins using a click-chemistry-based tagging

Gurcel, Caroline; Vercoutter-Edouart, Anne-Sophie; Fonbonne, Catherine; Mortuaire, Marlène; Salvador, Arnaud; Michalski, Jean-Claude; Lemoine, Jérôme
April 2008
Analytical & Bioanalytical Chemistry;Apr2008, Vol. 390 Issue 8, p2089
Academic Journal
The O-linked β- N-acetylglucosamine ( O-GlcNAc) modification is an abundant post-translational modification in eukaryotic cells. This dynamic glycosylation plays a fundamental role in the activity of many nuclear and cytoplasmic proteins and is associated with pathologies like type II diabetes, Alzheimer’s disease or some cancers. However the exact link between O-GlcNAc-modified proteins and their function in cells is largely undefined for most cases. Here we report a strategy based on the 1,3-dipolar cycloaddition, called click chemistry, between unnatural N-acetylglucosamine (GlcNAc) analogues (substituted with an azido or alkyne group) and the corresponding biotinylated probe to specifically detect, enrich and identify O-GlcNAc-modified proteins. This bio-orthogonal conjugation confirms that only azido analogue of GlcNAc is metabolized by the cell. Thanks to the biotin probe, affinity purification on streptavidin beads allowed us to identify 32 O-GlcNAc-azido-tagged proteins by LC-MS/MS analysis in an MCF-7 cellular model, 14 of which were previously unreported. This work illustrates the use of the click-chemistry-based strategy combined with a proteomic approach to get further insight into the pattern of O-GlcNAc-modified proteins and the biological significance of this post-translational modification. [Figure not available: see fulltext.]


Related Articles

  • Evanescent-field fluorescence-assisted lectin microarray: a new strategy for glycan profiling. Kuno, Atsushi; Uchiyama, Noboru; Koseki-Kuno, Shiori; Ebe, Youji; Takashima, Seigo; Yamada, Masao; Hirabayashi, Jun // Nature Methods;Nov2005, Vol. 2 Issue 11, p851 

    Glycans have important roles in living organisms with their structural diversity. Thus, glycomics, especially aspects involving the assignment of functional glycans in a high-throughput manner, has been an emerging field in the postproteomics era. To date, however, there has been no versatile...

  • Applications of Copper-Catalyzed Click Chemistry in Activity-Based Protein Profiling. Martell, Julianne; Weerapana, Eranthie // Molecules;Feb2014, Vol. 19 Issue 2, p1378 

    Activity-based protein profiling (ABPP) is a chemical proteomic technique that enables the interrogation of protein activity directly within complex proteomes. Given the dominant role of posttranslational modifications in regulating protein function in vivo, ABPP provides a direct readout of...

  • Analytical Technologies for Identification and Characterization of the Plant N-Glycoproteome. Ruiz-May, Eliel; Thannhauser, Theodore W.; Sheng Zhang; Rose, Jocelyn K. C. // Frontiers in Plant Science;Jun2012, Vol. 3, p1 

    N-glycosylation is one of the most common and complex post-translational modifications of eukaryotic proteins and one that has numerous roles, such as modulating protein stability, sorting, folding, enzyme activity, and ligand interactions. In plants, the functional significance of...

  • Synthesis of a Spiro D-Proline Analogue Bearing D-Fructose. Cipolla, Laura; Redaelli, Cristina; Nicotra, Francesco // Letters in Drug Design & Discovery;Jun2005, Vol. 2 Issue 4, p291 

    A bicyclic spiro analogue of D-proline was synthesized from perbenzylated methyl D-fructoside. The synthetic steps required first a three carbon chain elongation at the anomeric position by C-glycosylation, secondly the introduction of the amino function at C-1, and finally the cyclization to...

  • Cytoskeleton: Remodelling the FtsZ network. Du Toit, Andrea // Nature Reviews Molecular Cell Biology;Jan2014, Vol. 15 Issue 1, p3 

    The article highlights a study that examines Z-ring formation in vitro and uncover a dual role for FtsA in filamentation temperature-sensitive protein Z (FtsZ) network assembly and rearrangement. Fluorescently labelled FtsZ and FtsA proteins have been used by the researchers, and reconstituted...

  • Molecular mechanisms of imidazole and benzene ring binding in proteins. Zhuravlev, A. V.; Shchegolev, B. F.; Savvateeva-Popova, E. V.; Popov, A. V. // Biochemistry (00062979);Aug2009, Vol. 74 Issue 8, p925 

    Aromatic bonds of amino acid radicals play an important role in arrangement of protein primary structure. Previously, the existence of a number of preferable conformations of aromatic dimers was shown theoretically and experimentally, the best known of which are parallel-displaced and...

  • Introduction of an Artificial Cu Binding Site at the Surface of CA II: Pit-falls of Rational Design Finally Scooped by Serendipity. Wischeler, Johannes Schulze; Heine, Andreas; Klebe, Gerhard // Current Chemical Biology;Jan2011, Vol. 5 Issue 1, p1 

    The aim of this study was the introduction of an artificial copper center at the surface of carbonic anhydrase II (CA II) which subsequently might be used by azide and alkyne building blocks to catalyze triazole formation following 2+3 dipolar cycloaddition. Several mutants were produced to...

  • Asymmetric Synthesis of �-Lactams Through the Staudinger Reaction and Their Use as Building Blocks of Natural and Nonnatural Products. Palomo, C.; Aizpurua, J. M.; Ganboa, I.; Oiarbide, M. // Current Medicinal Chemistry;Jul2004, Vol. 11 Issue 14, p1837 

    In the last two decades, the better understanding of the mechanistic aspects of the �-lactams' biological activity and their inhibition, and the chemical exploitation of �-lactams as synthetic intermediates in organic chemistry, have experienced a continuous and somewhat complementary...

  • Protein binding: Antibody surrogates click into place. Kodadek, Thomas // Nature Chemistry;2009, Vol. 1 Issue 3, p183 

    The article presents a study conducted by Sharpless and colleagues which examines an alternative relying on target protein to guide a Huisgen cycloaddition between modest affinity ligands that carries an azide or alkyne. The study uses a high-affinity bivalent ligand for acetylcholinesterase...


Read the Article


Sorry, but this item is not currently available from your library.

Try another library?
Sign out of this library

Other Topics