PXD016041

PXD016041 is an original dataset announced via ProteomeXchange.

Title	selective protein O-glcNAcylation in cells by a proximity-directed O-GlcNAc transferase
Description	O-Linked N-acetylglucosamine (O-GlcNAc) is a monosaccharide that plays an essential role in cellular signaling throughout the nucleocytoplasmic proteome of eukaryotic cells. Yet, the study of post-translational modifications like O-GlcNAc has been limited by the lack of strategies to induce O-GlcNAcylation of a target protein in cells. Here, we report a generalizable genetic strategy to induce O-GlcNAc to specific target proteins in cells using a nanobody as a proximity-directing agent fused to O-GlcNAc transferase (OGT). Fusion of a nanobody that recognizes GFP (nGFP) or a nanobody that recognizes the four-amino acid sequence EPEA (nEPEA) to OGT(4), a truncated form of OGT, yielded a nanobody-OGT(4) construct that selectively delivered O-GlcNAc to the target protein (e.g., JunB, cJun, Nup62) and reduced alteration of global O-GlcNAc levels in the cell. Chemical proteomics confirmed the selective increase in O-GlcNAc to the target protein by nanobody-OGT(4). Glycoproteomics on the target protein revealed similar glycosite selectivity between nanobody-OGT(4) or global elevation of O-GlcNAc. Finally, we demonstrate the ability to selectively target endogenous ⍺-synuclein for glycosylation in HEK293T cells. Thus, the use of nanobodies to redirect OGT substrate selection is a versatile strategy to induce glycosylation to desired target proteins in cells that will facilitate discovery of O-GlcNAc functions and provide a mechanism to engineer O-GlcNAc signaling. The proximity-directed OGT approach for protein-selective O-GlcNAcylation is readily translated to additional protein targets and nanobodies, and may constitute a generalizable strategy to control post-translational modifications in cells.
HostingRepository	PRIDE
AnnounceDate	2024-10-22
AnnouncementXML	Submission_2024-10-22_05:31:36.680.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Christina Woo
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	No PTMs are included in the dataset
Instrument	LTQ Orbitrap

Revision	Datetime	Status	ChangeLog Entry
0	2019-10-28 02:27:40	ID requested
1	2022-02-15 09:46:37	announced
⏵ 2	2024-10-22 05:31:37	announced	2024-10-22: Updated project metadata.

Ramirez DH, Aonbangkhen C, Wu HY, Naftaly JA, Tang S, O'Meara TR, Woo CM, Engineering a Proximity-Directed O-GlcNAc Transferase for Selective Protein O-GlcNAcylation in Cells. ACS Chem Biol, 15(4):1059-1066(2020) [pubmed]

10.1021/acschembio.0c00074;

submitter keyword: OGT, glycoproteomics,nanobody, o-glcnac

Christina Woo
contact affiliation	Lab head
contact email	cwoo@chemistry.harvard.edu
lab head
Christina Woo
contact affiliation	Harvard University
contact email	cwoo@chemistry.harvard.edu
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2021/09/PXD016041

PRIDE project URI

• PRIDE
  1. PXD016041
     1. Label: PRIDE project
     2. Name: selective protein O-glcNAcylation in cells by a proximity-directed O-GlcNAc transferase