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A study of the structural properties of sites modified by the O-linked 6-N-acetylglucosamine transferase.

ABSTRACT: Protein O-GlcNAcylation (O-GlcNAc) is an essential post-translational modification (PTM) in higher eukaryotes. The O-linked ?-N-acetylglucosamine transferase (OGT), targets specific Serines and Threonines (S/T) in intracellular proteins. However, unlike phosphorylation, fewer than 25% of known O-GlcNAc sites match a clear sequence pattern. Accordingly, the three-dimensional structures of O-GlcNAc sites were characterised to investigate the role of structure in molecular recognition. From 1,584 O-GlcNAc sites in 620 proteins, 143 were mapped to protein structures determined by X-ray crystallography. The modified S/T were 1.7 times more likely to be annotated in the REM465 field which defines missing residues in a protein structure, while 7 O-GlcNAc sites were solvent inaccessible and unlikely to be targeted by OGT. 132 sites with complete backbone atoms clustered into 10 groups, but these were indistinguishable from clusters from unmodified S/T. This suggests there is no prevalent three-dimensional motif for OGT recognition. Predicted features from the 620 proteins were compared to unmodified S/T in O-GlcNAcylated proteins and globular proteins. The Jpred4 predicted secondary structure shows that modified S/T were more likely to be coils. 5/6 methods to predict intrinsic disorder indicated O-GlcNAcylated S/T to be significantly more disordered than unmodified S/T. Although the analysis did not find a pattern in the site three-dimensional structure, it revealed the residues around the modification site are likely to be disordered and suggests a potential role of secondary structure elements in OGT site recognition.

SUBMITTER: Britto-Borges T

PROVIDER: S-EPMC5590929 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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A study of the structural properties of sites modified by the O-linked 6-N-acetylglucosamine transferase.

Britto-Borges Thiago T Barton Geoffrey J GJ

PloS one 20170908 9

Protein O-GlcNAcylation (O-GlcNAc) is an essential post-translational modification (PTM) in higher eukaryotes. The O-linked β-N-acetylglucosamine transferase (OGT), targets specific Serines and Threonines (S/T) in intracellular proteins. However, unlike phosphorylation, fewer than 25% of known O-GlcNAc sites match a clear sequence pattern. Accordingly, the three-dimensional structures of O-GlcNAc sites were characterised to investigate the role of structure in molecular recognition. From 1,584 O ...[more]

PMID: 28886091

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Project description:BackgroundOverriding the differentiation blockage in acute myeloid leukemia (AML) is the most successful mode-of-action in leukemia therapy - now curing the vast majority of patients with acute promyelocytic leukemia (APL) using all-trans retinoic acid (ATRA)-based regimens. Similar approaches in other leukemia subtypes, such as IDH1/2-mutated AML, are under active investigation. We herein present successful release of the differentiation blockage upon treatment with the natural (-)-Δ9-Tetrahydrocannabinol isomer dronabinol in vitro and in vivo.MethodsCellular maturation and differentiation were followed in two patients employing whole genome methylation profiling, proteome analyses, NGS deep sequencing and multispectral imaging flow cytometry. For functional studies lentiviral OGT knock-down in vitro and ex vivo cell models were created to evaluate proliferative, apoptotic and differentiating effects of OGT in acute leukemia.FindingsIn here, we provide molecular evidence that dronbinol is capable to override the differentiation blockage of acute leukemia blasts at the state of the leukemia-initiating clone. We further identify the O-linked β-N-acetyl glucosamine (O-GlcNAc) transferase (OGT) to be crucial in this process. OGT is a master regulator enzyme adding O-GlcNAc to serine or threonine residues in a multitude of target proteins. Aberrant O-GlcNAc modification is implicated in pathologies of metabolic, neurodegenerative and autoimme diseases as well as cancers. We provide evidence that dronabinol induces transcription of OGT via epigenetic hypomethylation of the transcription start site (TSS). A lentiviral OGT-knock out approach proves the central role of OGT exerting antileukemic efficacy via a dual-mechanism of action: High concentrations of dronabinol result in induction of apoptosis, whereas lower concentrations drive cellular maturation. Most intriguingly, overriding of the differentiation blockage of acute leukemia blasts is validated in vivo following two patients treated with dronabinol.InterpretationIn conclusion, we provide evidence for overcoming the differentiation blockage in acute leukemia in subentities beyond promyelocytic and IDH1/2-mutated leukemia and thereby identify O-GlcNAcylation as a novel (drugable) field for future leukemia research.FundingUnrestricted grant support by the IZKF Program of the Medical Faculty Tübingen (MMS) and Brigitte Schlieben-Lange Program as well as the Margarete von Wrangell Program of the Ministry of Science, Research and the Arts, Baden-Württemberg, Germany (KKS) and Athene Program of the excellence initiative University of Tübingen (KKS).

Dataset Information

A study of the structural properties of sites modified by the O-linked 6-N-acetylglucosamine transferase.

Publications

A study of the structural properties of sites modified by the O-linked 6-N-acetylglucosamine transferase.

Similar Datasets

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