Genetic recoding to dissect the roles of site-specific protein O-GlcNAcylation

Andrii Gorelik; Sergio Galan Bartual; Vladimir Borodkin; Joby Varghese; Andrew Ferenbach; Daan van Aalten

doi:10.1038/s41594-019-0325-8

Genetic recoding to dissect the roles of site-specific protein O-GlcNAcylation

Andrii Gorelik, Sergio Galan Bartual, Vladimir Borodkin, Joby Varghese, Andrew Ferenbach, Daan van Aalten (Lead / Corresponding author)

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51 Citations (Scopus)

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Abstract

Modification of specific Ser and Thr residues of nucleocytoplasmic proteins with O-GlcNAc, catalyzed by O-GlcNAc transferase (OGT), is an abundant posttranslational event essential for proper animal development and is dysregulated in various diseases. Due to the rapid concurrent removal by the single O-GlcNAcase (OGA), precise functional dissection of site-specific O-GlcNAc modification in vivo is currently not possible without affecting the entire O-GlcNAc proteome. Exploiting the fortuitous promiscuity of OGT, we show that S-GlcNAc is a hydrolytically stable and accurate structural mimic of O-GlcNAc that can be encoded in mammalian systems with CRISPR-Cas9 in an otherwise unperturbed O-GlcNAcome. Using this approach, we target an elusive Ser 405 O-GlcNAc site on OGA, showing that this site-specific modification affects OGA stability.

Original language	English
Pages (from-to)	1071-1077
Number of pages	7
Journal	Nature Structural & Molecular Biology
Volume	26
Issue number	11
Early online date	6 Nov 2019
DOIs	https://doi.org/10.1038/s41594-019-0325-8
Publication status	Published - Nov 2019

ASJC Scopus subject areas

Structural Biology
Molecular Biology

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10.1038/s41594-019-0325-8

Author Accepted ManuscriptAccepted author manuscript, 5.16 MB

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@article{184822540fc1455eba581c38f499229d,

title = "Genetic recoding to dissect the roles of site-specific protein O-GlcNAcylation",

abstract = "Modification of specific Ser and Thr residues of nucleocytoplasmic proteins with O-GlcNAc, catalyzed by O-GlcNAc transferase (OGT), is an abundant posttranslational event essential for proper animal development and is dysregulated in various diseases. Due to the rapid concurrent removal by the single O-GlcNAcase (OGA), precise functional dissection of site-specific O-GlcNAc modification in vivo is currently not possible without affecting the entire O-GlcNAc proteome. Exploiting the fortuitous promiscuity of OGT, we show that S-GlcNAc is a hydrolytically stable and accurate structural mimic of O-GlcNAc that can be encoded in mammalian systems with CRISPR-Cas9 in an otherwise unperturbed O-GlcNAcome. Using this approach, we target an elusive Ser 405 O-GlcNAc site on OGA, showing that this site-specific modification affects OGA stability.",

author = "Andrii Gorelik and {Galan Bartual}, Sergio and Vladimir Borodkin and Joby Varghese and Andrew Ferenbach and {van Aalten}, Daan",

note = "This work was funded by a Wellcome Trust Investigator Award (110061) to DvA and a Wellcome Trust 4-year PhD studentship (105310/Z/14/Z) to AG. ",

year = "2019",

month = nov,

doi = "10.1038/s41594-019-0325-8",

language = "English",

volume = "26",

pages = "1071--1077",

journal = "Nature Structural & Molecular Biology",

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AU - Gorelik, Andrii

AU - Galan Bartual, Sergio

AU - Borodkin, Vladimir

AU - Varghese, Joby

AU - Ferenbach, Andrew

AU - van Aalten, Daan

N1 - This work was funded by a Wellcome Trust Investigator Award (110061) to DvA and a Wellcome Trust 4-year PhD studentship (105310/Z/14/Z) to AG.

PY - 2019/11

Y1 - 2019/11

N2 - Modification of specific Ser and Thr residues of nucleocytoplasmic proteins with O-GlcNAc, catalyzed by O-GlcNAc transferase (OGT), is an abundant posttranslational event essential for proper animal development and is dysregulated in various diseases. Due to the rapid concurrent removal by the single O-GlcNAcase (OGA), precise functional dissection of site-specific O-GlcNAc modification in vivo is currently not possible without affecting the entire O-GlcNAc proteome. Exploiting the fortuitous promiscuity of OGT, we show that S-GlcNAc is a hydrolytically stable and accurate structural mimic of O-GlcNAc that can be encoded in mammalian systems with CRISPR-Cas9 in an otherwise unperturbed O-GlcNAcome. Using this approach, we target an elusive Ser 405 O-GlcNAc site on OGA, showing that this site-specific modification affects OGA stability.

AB - Modification of specific Ser and Thr residues of nucleocytoplasmic proteins with O-GlcNAc, catalyzed by O-GlcNAc transferase (OGT), is an abundant posttranslational event essential for proper animal development and is dysregulated in various diseases. Due to the rapid concurrent removal by the single O-GlcNAcase (OGA), precise functional dissection of site-specific O-GlcNAc modification in vivo is currently not possible without affecting the entire O-GlcNAc proteome. Exploiting the fortuitous promiscuity of OGT, we show that S-GlcNAc is a hydrolytically stable and accurate structural mimic of O-GlcNAc that can be encoded in mammalian systems with CRISPR-Cas9 in an otherwise unperturbed O-GlcNAcome. Using this approach, we target an elusive Ser 405 O-GlcNAc site on OGA, showing that this site-specific modification affects OGA stability.

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DO - 10.1038/s41594-019-0325-8

M3 - Article

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JO - Nature Structural & Molecular Biology

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Genetic recoding to dissect the roles of site-specific protein O-GlcNAcylation

Abstract

ASJC Scopus subject areas

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Genetic and Chemical Validation of Sugar Nucleotide Biosynthesis as a Target Against Candida Albicans (Joint with University of Aberdeen)

Molecular Mechanisms of O-GICNAC Signalling (Investigator award)

Molecular and Cellular Biology 4 Year PhD

Cite this

Genetic recoding to dissect the roles of site-specific protein O-GlcNAcylation

Abstract

ASJC Scopus subject areas

Access to Document

Other files and links

Fingerprint

Projects

Genetic and Chemical Validation of Sugar Nucleotide Biosynthesis as a Target Against Candida Albicans (Joint with University of Aberdeen)

Molecular Mechanisms of O-GICNAC Signalling (Investigator award)

Molecular and Cellular Biology 4 Year PhD

Cite this