Endo-M Mediated Chemoenzymatic Approach Enables Reversible Glycopeptide Labeling for O-GlcNAcylation Analysis

Yao Chen; Feng Tang; Hongqiang Qin; Xuyang Yue; Yongzhan Nie; Wei Huang; Mingliang Ye

doi:10.1002/anie.202117849

Endo-M Mediated Chemoenzymatic Approach Enables Reversible Glycopeptide Labeling for O-GlcNAcylation Analysis

Angew Chem Int Ed Engl. 2022 Jun 7;61(23):e202117849. doi: 10.1002/anie.202117849. Epub 2022 Mar 30.

Authors

Yao Chen^{1

2}, Feng Tang^{2

3}, Hongqiang Qin^{1

2}, Xuyang Yue¹, Yongzhan Nie⁴, Wei Huang^{2

3}, Mingliang Ye^{1

2}

Affiliations

¹ CAS Key Laboratory of Separation Science for Analytical Chemistry, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, 116023, Dalian, China.
² University of Chinese Academy of Sciences, 101408, Beijing, China.
³ CAS Key Laboratory of Receptor Research, CAS Center for Excellence in Molecular Cell Science, Center for Biotherapeutics Discovery Research, Shanghai Institute of Material Medical, Chinese Academy of Sciences, 201203, Shanghai, China.
⁴ State Key Laboratory of Cancer Biology, National Clinical Research Center for Digestive Diseases and Xijing Hospital of Digestive Diseases, Fourth Military Medical University, 710032, Xi'an, China.

PMID: 35289036
DOI: 10.1002/anie.202117849

Abstract

To selectively enrich O-linked β-N-acetylglucosamine (O-GlcNAc) peptides in their original form from complex samples, we report the first reversible chemoenzymatic labeling approach for proteomic analysis. In this strategy, the O-GlcNAc moieties are ligated with long N-glycans using an Endo-M mutant, which enables the enrichment of the labeled glycopeptides by hydrophilic interaction liquid chromatography (HILIC). The attached glycans on the enriched glycopeptides are removed by wild-type Endo-M/S to restore the O-GlcNAc moiety. Compared with classic chemoenzymatic labeling, this approach enables the tag-free identification, and eliminates the interference of bulky tags in glycopeptide detection. This approach presents a unique avenue for the proteome-wide analysis of protein O-GlcNAcylation to promote its mechanism research.

Keywords: Chemoenzymatic Labeling; Glycopeptide Enrichment; Glycoproteomics; Hydrophilic Interaction Liquid Chromatography; O-GlcNAcylation.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Acetylglucosamine / metabolism
Chromatography, Liquid / methods
Glycopeptides* / chemistry
Polysaccharides / chemistry
Proteome / analysis
Proteomics*

Substances

Glycopeptides
Polysaccharides
Proteome
Acetylglucosamine