기술이전

 

탄뎀 질량분석기를 이용한 혈액 당단백질내 알파갈락토실화의 선택적 검출법

관리번호
RS05202012150001
첨부파일
개발년도
2020
저자정보
유종신
참고문헌
Anal. Chem. 2020, 92, 13144−13154
관련장비
초고분해능 융합질량분석기
응용분야
1. 바이오의약품 품질검사 / 2. 혈액 당단백질의 면역원성 규명
연구분야
BT
분석/적용분야
의약품분석
기술분야
질량분석
분석/공동연구지원
1. 인간 혈액내 당단백질 분석 2. 마우스 혈액내 당단백질 분석
요약
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).
키워드
α‑Galactosyl Epitopes, N‑Glycoproteins, Fragment Ions, Higher-Energy Collisional Dissociation
연구개발성과
1. 인간 혈액내 당단백질 구조분석 2. 유전자 변형 마우스의 당단백질 특성분석
내 용
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 αgalactosyl 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 Nglycoproteins 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 Nglycopeptides. 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 Nglycopeptides 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.