2. Academic Validation
  3. Development of derivatization-enhanced EIEIO-driven glycosidic linkage sequencing (DEED-GL-Seq) strategy and its application to glycogen-sourced oligosaccharides profiling

Development of derivatization-enhanced EIEIO-driven glycosidic linkage sequencing (DEED-GL-Seq) strategy and its application to glycogen-sourced oligosaccharides profiling

  • Carbohydr Polym. 2025 Oct 15:366:123877. doi: 10.1016/j.carbpol.2025.123877.
Xin Zheng 1,  Yufang Ma 1,  Xinge Cui 2,  Yi Wang 3,  Jingjing Jiang 4,  Ting Liu 5,  Zhijun Zhang 6,  Mingsheng Ma 7,  Xiaohong Han 8,  Cai Tie 9
Affiliations

Affiliations

  • 1 Clinical Pharmacology Research Center, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100730, China; NMPA Key Laboratory for Clinical Research and Evaluation of Drug, Beijing Key Laboratory of Key Technologies for Early Clinical Trial Evaluation of Innovative Drugs for Major Diseases, Beijing 100730, China.
  • 2 Institute of Clinical Medicine, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100730, China.
  • 3 State Key Laboratory for Fine Exploration and Intelligent Development of Coal Resources, School of Chemical and Environmental Engineering, China University of Mining and Technology-Beijing, Ding11 Xueyuan Road, Beijing 100083, China.
  • 4 Department of Pediatrics, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100730, China.
  • 5 SCIEX, Analytical Instrument Trading Co., Ltd., Shanghai 200335, China.
  • 6 School of Chemical and Environmental Engineering, China University of Mining and Technology-Beijing, Beijing 100083, China.
  • 7 Department of Pediatrics, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100730, China. Electronic address: mamingsheng@pumch.cn.
  • 8 Clinical Pharmacology Research Center, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100730, China; NMPA Key Laboratory for Clinical Research and Evaluation of Drug, Beijing Key Laboratory of Key Technologies for Early Clinical Trial Evaluation of Innovative Drugs for Major Diseases, Beijing 100730, China. Electronic address: Hanxiaohong@pumch.cn.
  • 9 State Key Laboratory for Fine Exploration and Intelligent Development of Coal Resources, School of Chemical and Environmental Engineering, China University of Mining and Technology-Beijing, Ding11 Xueyuan Road, Beijing 100083, China. Electronic address: tiecai@cumtb.edu.cn.
PMID: 40733807 DOI: 10.1016/j.carbpol.2025.123877
Abstract

The structural complexity arising from diverse glycosidic linkages in oligosaccharides hinders the elucidation of their biological functions. Existing analytical methods often lack the necessary feasibility and accessibility for comprehensive linkage analysis. To address this, we developed Derivatization-Enhanced EIEIO-Driven Glycosidic Linkage Sequencing (DEED-GL-Seq), a novel strategy leveraging differential electron density modulation around glycosidic bonds upon N2,N2,N4,N4-tetraethyl-6-hydrazineyl-1,3,5-triazine-2,4-diamine (T3) derivatization. This method integrates EIEIO MS2 to identify glycosidic bond-specific diagnostic fragments for linkage determination. Validation confirmed DEED-GL-Seq's specificity, sensitivity, reliability, and standard-independence. Coupling with fine-tuned HILIC separation further enhanced its analytical power for complex matrices. We applied DEED-GL-Seq to profile oligosaccharides derived from partially hydrolyzed glycogen, validating our results against reference standards. By analyzing the abundance ratios of specific trisaccharides, we predicted the branching degrees of glycogen and amylopectin, corroborating existing literature. Notably, DEED-GL-Seq identified established (Glc₄) and novel (HEX-1, HEX-2, HEPTA-1, HEPTA-2) potential glycosidic biomarkers for GSD-II in urine, with the novel oligosaccharides showing superior diagnostic performance. The unique structure of HEX-2 suggests distinct biosynthetic pathways in GSD-II pathophysiology. DEED-GL-Seq represents a significant advancement in glycomics, offering a powerful tool for comprehensive oligosaccharide profiling and laying a foundation for in-depth functional investigations. This work presents a novel application paradigm for EIEIO technology.

Keywords

EIEIO; GSD-II biomarker; Glycogen; Glycosidic linkages sequencing; Oligosaccharides profiling.

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