Human O-GlcNAcase binds substrates in a conserved peptide recognition groove

Marianne Schimpl , Alexander Wolfgang Schuettelkopf , Vladimir S Borodkin , Daan Mf van Aalten (1)

1 Wellcome Trust Biocentre

Modification of cellular proteins with N-acetylglucosamine (O-GlcNAc) competes with protein phosphorylation and regulates a plethora of cellular processes. O-GlcNAcylation is orchestrated by two opposing enzymes, O-GlcNAc transferase and O-GlcNAcase, which recognize their target proteins via as yet unidentified mechanisms. Here, we uncover the first insights into the mechanism of substrate recognition by human O-GlcNAcase. The structure of a novel bacterial O-GlcNAcase orthologue reveals a putative substrate binding groove, conserved with metazoan O-GlcNAcases. Guided by this structure, conserved amino acids lining this groove in human O-GlcNAcase were mutated and the activity on three different substrate proteins (TAB1, FoxO1 and CREB) was tested in an in vitro de-glycosylation assay. The results provide the first evidence that human O-GlcNAcase may possess a substrate recognition mechanism that involves interactions with O-GlcNAcylated proteins beyond the GlcNAc binding site, with possible implications for differential regulation of cycling of O-GlcNAc on different proteins.

Mots clés: Life Sciences

Marianne Schimpl, Alexander Wolfgang Schuettelkopf, Vladimir S Borodkin, Daan Mf van Aalten. Human O-GlcNAcase binds substrates in a conserved peptide recognition groove. Biochemical Journal, 2010, 432 (1), pp.1-7. ⟨10.1042/BJ20101338⟩. ⟨hal-00529111⟩