Proteomics

Dataset Information

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Targeted deglycosylation with a nanobody-directed split OGA in living cells


ABSTRACT: O-GlcNAc is an essential and dynamic post-translational modification present on thousands of nucleocytoplasmic proteins. Interrogating the role of O-GlcNAc on a target protein in cells is critical yet challenging without disturbing O-GlcNAc globally or after extensive glycosite mapping. Herein, we developed a nanobody-fused split O-GlcNAcase (OGA) as a targeted O-GlcNAc eraser for protein-selective O-GlcNAc removal in cells. Through systematic screening, we identified the essential domains of OGA and afforded a split OGA with limited activity, which selectively reinstated deglycosylation activity on the target protein upon fusion of a nanobody in living cells. We demonstrate the generality of the O-GlcNAc eraser against a series of target proteins and reveal that O-GlcNAc stabilizes the transcription factor c-Jun and promotes its interaction with c-Fos. Thus, nanobody-directed split OGA speeds the selective removal and functional evaluation of O-GlcNAc on individual proteins via an approach that is generalizable to a broader array of post-translational modifications.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): T Cell, Kidney

SUBMITTER: Christina Woo  

LAB HEAD: Christina Woo

PROVIDER: PXD018914 | Pride | 2022-02-15

REPOSITORIES: Pride

Dataset's files

Source:
Action DRS
190930L_SAM6100_YG147_TF_TMT_final no normal.pride.mgf.gz Mgf
190930L_SAM6100_YG147_TF_TMT_finalnonormal.mzML Mzml
Yun_OGA_mzid.mzid.gz Mzid
Yun_OGA_mzid.pride.mztab.gz Mztab
Yun_OGA_rawdata.raw Raw
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Publications

Target protein deglycosylation in living cells by a nanobody-fused split O-GlcNAcase.

Ge Yun Y   Ramirez Daniel H DH   Yang Bo B   D'Souza Alexandria K AK   Aonbangkhen Chanat C   Wong Stephanie S   Woo Christina M CM  

Nature chemical biology 20210308 5


O-linked N-acetylglucosamine (O-GlcNAc) is an essential and dynamic post-translational modification that is presented on thousands of nucleocytoplasmic proteins. Interrogating the role of O-GlcNAc on a single target protein is crucial, yet challenging to perform in cells. Herein, we developed a nanobody-fused split O-GlcNAcase (OGA) as an O-GlcNAc eraser for selective deglycosylation of a target protein in cells. After systematic cellular optimization, we identified a split OGA with reduced inhe  ...[more]

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