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A rapid and efficient chemical strategy has been developed to identify proteins with a mysterious modification--the attachment of oxygen-linked -N-acetylglucosamine (O-GlcNAc) sugars. GLYCOPROBES Azides introduced into O-GlcNAc-modified proteins serve as chemical handles for introducing detectable probes. ADAPTED WITH PERMISSION FROM PNAS O-GlcNAc modification occurs widely on proteins in the nucleus and cytoplasm of multicellular plants and animals. The function of this glycosylation process is poorly understood, but it may play a key role in transcription, cytoskeletal organization, and signaling, and O-GlcNAc metabolism may be abnormally regulated in diabetes. Tools have been developed recently to aid identification of O-GlcNAc-modified proteins, including site-specific antibodies and a liquid chromatography-tandem mass spectrometry technique. Now, chemistry professor Carolyn R. Bertozzi of Howard Hughes Medical Institute and the University of California, Berkeley, postdoctoral fellow David J. Vocadlo, and coworkers have devised a complementary approach that may prove more useful for high-throughput proteomics applications [Proc. Natl. Acad. Sci. USA, published online July 21, http://www.pnas.org/cgi/content/abstract/ 1632821100v1]. "We found that O-GlcNAc residues on cellular proteins can be metabolically labeled with azido groups, which primes them for chemical modification via the Staudinger ligation, a reaction we developed a few years back for specific bioconjugation," Bertozzi explains. "Using this chemistry on whole cell lysates, we were able to discriminate proteins modified with O-GlcNAc from the rest of the proteome--setting the stage for an inventory of the complete repertoire of O-GlcNAc-modified proteins." Biological chemistry professor Gerald W. Hart of Johns Hopkins University School of Medicine points out some drawbacks of the technique: It modifies only a small percentage of O-GlcNAc groups at any one site on a protein, and the required reagents are not available commercially. "Nonetheless, for many types of studies, it should prove a valuable method," he says |
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