Enzymatic addition of O-GlcNAc to nuclear and cytoplasmic proteins. Identification of a uridine diphospho-N-acetylglucosamine:peptide beta-N-acetylglucosaminyltransferase - PubMed (original) (raw)
. 1990 Feb 15;265(5):2563-8.
Affiliations
- PMID: 2137449
Free article
Enzymatic addition of O-GlcNAc to nuclear and cytoplasmic proteins. Identification of a uridine diphospho-N-acetylglucosamine:peptide beta-N-acetylglucosaminyltransferase
R S Haltiwanger et al. J Biol Chem. 1990.
Free article
Abstract
An assay for the enzyme responsible for the addition of O-linked N-acetylglucosamine (O-GlcNAc) to proteins, a UDP-N-acetylglucosamine:peptide N-acetylglucosaminyltransferase, is reported using the synthetic peptide YSDSPSTST as the acceptor substrate. The activity is linearly dependent on time, enzyme, and substrate concentration. Replacement of the proline with a glycine in the peptide renders it ineffective as a substrate, whereas changing of the aspartic acid to a glycine has no effect. Product characterization of the glycosylated peptide demonstrates that the monosaccharide covalently attached to the peptide is N-acetylglucosamine (GlcNAc) and has not been epimerized to N-acetylgalactosamine. Mild base-catalyzed beta-elimination of the in vitro glycosylated peptide quantitatively yields GlcNAcitol, indicating that the GlcNAc is attached via an O-linkage. The transferase activity is strongly inhibited by UDP but is unaffected by GlcNAc or tunicamycin. Interestingly, EDTA only slightly inhibits activity, suggesting that the enzyme may not require divalent cations. The majority of the activity is soluble, and the remainder is lost from membranes after extracting with high salt and EDTA. Consistent with the subcellular localization of most proteins bearing O-GlcNAc, the activity appears to reside in the cytosolic portion of the cell when compared to two lumenal marker enzymes, galactosyltransferase and mannose-6-phosphatase.
Similar articles
- E. coli sabotages the in vivo production of O-linked β-N-acetylglucosamine-modified proteins.
Goodwin OY, Thomasson MS, Lin AJ, Sweeney MM, Macnaughtan MA. Goodwin OY, et al. J Biotechnol. 2013 Dec;168(4):315-23. doi: 10.1016/j.jbiotec.2013.10.008. Epub 2013 Oct 16. J Biotechnol. 2013. PMID: 24140293 - Molecular mechanisms regulating O-linked N-acetylglucosamine (O-GlcNAc)-processing enzymes.
King DT, Males A, Davies GJ, Vocadlo DJ. King DT, et al. Curr Opin Chem Biol. 2019 Dec;53:131-144. doi: 10.1016/j.cbpa.2019.09.001. Epub 2019 Oct 22. Curr Opin Chem Biol. 2019. PMID: 31654859 Review.
Cited by
- O-GlcNAcylation promotes malignancy and cisplatin resistance of lung cancer by stabilising NRF2.
Zhang Y, Sun C, Ma L, Xiao G, Gu Y, Yu W. Zhang Y, et al. Clin Transl Med. 2024 Oct;14(10):e70037. doi: 10.1002/ctm2.70037. Clin Transl Med. 2024. PMID: 39358921 Free PMC article. - O-GlcNAc informatics: advances and trends.
Hou C, Li W, Li Y, Ma J. Hou C, et al. Anal Bioanal Chem. 2024 Sep 18. doi: 10.1007/s00216-024-05531-2. Online ahead of print. Anal Bioanal Chem. 2024. PMID: 39294469 Review. - Identification of a Polypeptide Inhibitor of _O_-GlcNAc Transferase with Picomolar Affinity.
Hammel FA, Payne NC, Marando VM, Mazitschek R, Walker S. Hammel FA, et al. J Am Chem Soc. 2024 Sep 25;146(38):26320-26330. doi: 10.1021/jacs.4c08656. Epub 2024 Sep 14. J Am Chem Soc. 2024. PMID: 39276112 Free PMC article. - High aggressiveness of papillary thyroid cancer: from clinical evidence to regulatory cellular networks.
Zhang J, Xu S. Zhang J, et al. Cell Death Discov. 2024 Aug 26;10(1):378. doi: 10.1038/s41420-024-02157-2. Cell Death Discov. 2024. PMID: 39187514 Free PMC article. Review. - A protein O-GlcNAc glycosyltransferase regulates the antioxidative response in Yersinia pestis.
Cao S, Wang T, Ren Y, Wu G, Zhang Y, Tan Y, Zhou Y, Chen H, Zhang Y, Song Y, Yang R, Du Z. Cao S, et al. Nat Commun. 2024 Aug 16;15(1):7062. doi: 10.1038/s41467-024-50959-w. Nat Commun. 2024. PMID: 39152136 Free PMC article.
Publication types
MeSH terms
Substances
LinkOut - more resources
Full Text Sources
Other Literature Sources
Molecular Biology Databases