Selective Identification of α-Galactosyl Epitopes in N-Glycoproteins Using Characteristic Fragment Ions from Higher-Energy Collisional Dissociation
| Autor: | Yong-Sam Kim, Jeong Heon Ko, Gun Wook Park, Jong Hwan Shin, Hyun Joo An, Ju Yeon Lee, Dae-In Ha, Jong Shin Yoo, Hyun Kyoung Lee, Jin Young Kim, Jeong Gu Kang, Kun Cho, Su Bin Moon |
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| Rok vydání: | 2020 |
| Předmět: |
chemistry.chemical_classification
Glycan Molecular mass biology Stereochemistry 010401 analytical chemistry 010402 general chemistry Mass spectrometry Tandem mass spectrometry 01 natural sciences Dissociation (chemistry) Epitope 0104 chemical sciences Analytical Chemistry carbohydrates (lipids) chemistry Antigen biology.protein lipids (amino acids peptides and proteins) Glycoprotein |
| Zdroj: | Analytical Chemistry. 92:13144-13154 |
| ISSN: | 1520-6882 0003-2700 |
| DOI: | 10.1021/acs.analchem.0c02276 |
| Popis: | The α-galactosyl epitope is a terminal N-glycan moiety of glycoproteins found in mammals except in humans, and thus, it is recognized as an antigen that provokes an immunogenic response in humans. Accordingly, it is necessary to analyze the α-galactosyl structure in biopharmaceuticals or organ transplants. Due to an identical glycan composition and molecular mass between α-galactosyl N-glycans and hybrid/high-mannose-type N-glycans, it is challenging to characterize α-galactosyl epitopes in N-glycoproteins using mass spectrometry. Here, we describe a method to identify α-galactosyl N-glycopeptides in mice glycoproteins using liquid chromatography with tandem mass spectrometry with higher-energy collisional dissociation (HCD). The first measure was an absence of [YHM] ion peaks in the HCD spectra, which was exclusively observed in hybrid and/or high-mannose-type N-glycopeptides. The second complementary criterion was the ratio of an m/z 528.19 (Hex2HexNAc1) ion to m/z 366.14 (Hex1HexNAc1) ion (Im/z528/Im/z366). The measure of [Im/z528/Im/z366 > 0.3] enabled a clear-cut determination of α-galactosyl N-glycopeptides with high accuracy. In Ggta1 knockout mice, we could not find any α-galactosyl N-glycoproteins identified in WT mice plasma. Using this method, we could screen for α-galactosyl N-glycoproteins from mice spleen, lungs, and plasma samples in a highly sensitive and specific manner. Conclusively, we suggest that this method will provide a robust analytical tool for determination of α-galactosyl epitopes in pharmaceuticals and complex biological samples. |
| Databáze: | OpenAIRE |
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