Structural insights into mechanism and specificity of O-GlcNAc transferase

Andrew J. Clarke, Ramon Hurtado-Guerrero, Shalini Pathak, Alexander Schuettelkopf, Vladimir Borodkin, Sharon M. Shepherd, Adel F. M. Ibrahim, Daan M. F. van Aalten (Lead / Corresponding author)

    Research output: Contribution to journalArticlepeer-review

    96 Citations (Scopus)


    Post-translational modification of protein serines/threonines with N-acetylglucosamine (O-GlcNAc) is dynamic, inducible and abundant, regulating many cellular processes by interfering with protein phosphorylation. O-GlcNAcylation is regulated by O-GlcNAc transferase (OGT) and O-GlcNAcase, both encoded by single, essential, genes in metazoan genomes. It is not understood how OGT recognises its sugar nucleotide donor and performs O-GlcNAc transfer onto proteins/peptides, and how the enzyme recognises specific cellular protein substrates. Here, we show, by X-ray crystallography and mutagenesis, that OGT adopts the (metal-independent) GT-B fold and binds a UDP-GlcNAc analogue at the bottom of a highly conserved putative peptide-binding groove, covered by a mobile loop. Strikingly, the tetratricopeptide repeats (TPRs) tightly interact with the active site to form a continuous 120A putative interaction surface, whereas the previously predicted phosphatidylinositide-binding site locates to the opposite end of the catalytic domain. On the basis of the structure, we identify truncation/point mutants of the TPRs that have differential effects on activity towards proteins/peptides, giving first insights into how OGT may recognise its substrates.

    Original languageEnglish
    Pages (from-to)2780-2788
    Number of pages9
    JournalEMBO Journal
    Issue number20
    Early online date25 Sept 2008
    Publication statusPublished - 22 Oct 2008


    • Glycobiology
    • O-GlcNAc
    • Protein structure
    • Signal transduction
    • Linked GlcNAc
    • N-acetylglucosamine deacetylase
    • Tetratricopeptide repeats
    • UDP-GlcNAc
    • Substrate specificity
    • Protein phosphatase
    • Mutational analysis
    • Cytosolic proteins
    • Insulin resistance
    • Crystal structures


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