A Chemoenzymatic Approach toward the Rapid and Sensitive Detection of O-GlcNAc Posttranslational Modifications

We report a new chemoenzymatic strategy for the rapid and sensitive detection of O-GlcNAc posttranslational modifications. The approach exploits the ability of an engineered mutant of β-1,4-galactosyltransferase to selectively transfer an unnatural ketone functionality onto O-GlcNAc glycosylated pro...

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Bibliographic Details
Published in:Journal of the American Chemical Society 2003-12, Vol.125 (52), p.16162-16163
Main Authors: Khidekel, Nelly, Arndt, Sabine, Lamarre-Vincent, Nathan, Lippert, Alexander, Poulin-Kerstien, Katherine G, Ramakrishnan, Boopathy, Qasba, Pradman K, Hsieh-Wilson, Linda C
Format: Article
Language:English
Subjects:
Acetylglucosamine - analysis
Acetylglucosamine - metabolism
alpha-Crystallins - analysis
alpha-Crystallins - metabolism
Animals
Bacterial Proteins - chemistry
Bacterial Proteins - metabolism
Biotin
Cyclic AMP Response Element-Binding Protein - analysis
Cyclic AMP Response Element-Binding Protein - metabolism
Glycoproteins - analysis
Glycoproteins - metabolism
Glycosylation
Horseradish Peroxidase - chemistry
Horseradish Peroxidase - metabolism
Luminescent Measurements
N-Acetyllactosamine Synthase - chemistry
N-Acetyllactosamine Synthase - metabolism
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Summary:We report a new chemoenzymatic strategy for the rapid and sensitive detection of O-GlcNAc posttranslational modifications. The approach exploits the ability of an engineered mutant of β-1,4-galactosyltransferase to selectively transfer an unnatural ketone functionality onto O-GlcNAc glycosylated proteins. Once transferred, the ketone moiety serves as a versatile handle for the attachment of biotin, thereby enabling chemiluminescent detection of the modified protein. Importantly, this approach permits the rapid visualization of proteins that are at the limits of detection using traditional methods. Moreover, it bypasses the need for radioactive precursors and captures the glycosylated species without perturbing metabolic pathways. We anticipate that this general chemoenzymatic strategy will have broad application to the study of posttranslational modifications.
ISSN:0002-7863
1520-5126
DOI:10.1021/ja038545r