Protocol for automated N-glycan sequencing using mass spectrometry and computer-assisted intelligent fragmentation

Chuncui Huang; Hui Wang; Jinyu Zhou; Yikang Huang; Yihui Ren; Keli Zhao; Yaojun Wang; Meijie Hou; Jingwei Zhang; Yaming Liu; Xinyue Ma; Jingyu Yan; Dongbo Bu; Wengang Chai; Shiwei Sun; Yan Li

doi:10.1016/j.xpro.2024.102976

Protocol for automated N-glycan sequencing using mass spectrometry and computer-assisted intelligent fragmentation

STAR Protoc. 2024 Jun 21;5(2):102976. doi: 10.1016/j.xpro.2024.102976. Epub 2024 Apr 16.

Authors

Chuncui Huang¹, Hui Wang², Jinyu Zhou³, Yikang Huang⁴, Yihui Ren⁴, Keli Zhao⁵, Yaojun Wang⁶, Meijie Hou⁷, Jingwei Zhang⁷, Yaming Liu³, Xinyue Ma⁸, Jingyu Yan⁹, Dongbo Bu⁴, Wengang Chai¹⁰, Shiwei Sun¹¹, Yan Li¹²

Affiliations

¹ Key Laboratory of Epigenetic Regulation and Intervention, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Beijing 100101, China; Western Institute of Health Data Science, 28 High Tech Avenue, Chongqing 401329, China.
² Division of Life Science and State Key Laboratory of Molecular Neuroscience, The Hong Kong University of Science and Technology, Clear Water Bay Ne Laboratory, Kowloon, Hong Kong SAR, China.
³ Key Laboratory of Epigenetic Regulation and Intervention, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Beijing 100101, China.
⁴ Key Laboratory of Intelligent Information Processing, Institute of Computing Technology, Chinese Academy of Sciences, 6 Kexueyuan South Road, Beijing 100080, China; University of Chinese Academy of Sciences, 19 Yuquan Road, Beijing 100049, China.
⁵ Key Laboratory of Intelligent Information Processing, Institute of Computing Technology, Chinese Academy of Sciences, 6 Kexueyuan South Road, Beijing 100080, China; Western Institute of Health Data Science, 28 High Tech Avenue, Chongqing 401329, China.
⁶ College of Information and Electrical Engineering, China Agricultural University, Beijing 100083, China.
⁷ Key Laboratory of Intelligent Information Processing, Institute of Computing Technology, Chinese Academy of Sciences, 6 Kexueyuan South Road, Beijing 100080, China.
⁸ Key Laboratory of Epigenetic Regulation and Intervention, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Beijing 100101, China; University of Chinese Academy of Sciences, 19 Yuquan Road, Beijing 100049, China.
⁹ Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Key Laboratory of Separation Science for Analytical Chemistry, Dalian 116023, China.
¹⁰ Glycosciences Laboratory, Department of Medicine, Imperial College London, W120NN London, UK. Electronic address: [email protected].
¹¹ Key Laboratory of Intelligent Information Processing, Institute of Computing Technology, Chinese Academy of Sciences, 6 Kexueyuan South Road, Beijing 100080, China; University of Chinese Academy of Sciences, 19 Yuquan Road, Beijing 100049, China. Electronic address: [email protected].
¹² Key Laboratory of Epigenetic Regulation and Intervention, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Beijing 100101, China; University of Chinese Academy of Sciences, 19 Yuquan Road, Beijing 100049, China; Western Institute of Health Data Science, 28 High Tech Avenue, Chongqing 401329, China. Electronic address: [email protected].

Abstract

Biological functions of glycans are intimately linked to fine details in branches and linkages, which make structural identification extremely challenging. Here, we present a protocol for automated N-glycan sequencing using multi-stage mass spectrometry (MSⁿ). We describe steps for release/purification and derivation of glycans and procedures for MSⁿ scanning. We then detail "glycan intelligent precursor selection" to computationally guide MSⁿ experiments. The protocol can be used for both discrete individual glycans and isomeric glycan mixtures. For complete details on the use and execution of this protocol, please refer to Sun et al.,¹ Huang et al.,² and Huang et al.³.

Keywords: Bioinformatics; Biotechnology and bioengineering; Chemistry; Computer sciences; Protein Biochemistry.

MeSH terms

Mass Spectrometry* / methods
Polysaccharides* / analysis
Polysaccharides* / chemistry
Sequence Analysis / methods

Substances

Polysaccharides

Grants and funding

WT_/Wellcome Trust/United Kingdom