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Reversed-phase depletion coupled with hydrophilic affinity enrichment for the selective isolation of N-linked glycopeptides by using Click OEG-CD matrix

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Abstract

Selective enrichment of glycopeptides is of great importance for protein glycosylation analysis using mass spectrometry since the signals of glycopeptides could be severely suppressed by the coexisting non-glycosylated peptides in the protein digest. In the present work, a strategy for N-linked glycopeptide enrichment through reversed-phase depletion coupled with hydrophilic affinity enrichment by applying the customized matrix named Click OEG-CD is developed. Compared with single hydrophilic interaction liquid chromatography (HILIC) mode, the strategy exhibited remarkably higher selectivity for N-linked glycopeptides. As many as 22, 18, and eight glycopeptides were detected in the glycopeptide fraction enriched with the strategy from the digests of human immunoglobulin G, horseradish peroxidase and bovine ribonuclease B, respectively. In addition, the strategy also showed high glycosylation microheterogeneity coverage for the enrichment of human α1-acid glycoprotein glycopeptides. More than 170 glycopeptides covering all the glycosylation sites were detected in the enriched fraction. The revered-phase liquid chromatography depletion coupled with HILIC enrichment strategy by using Click OEG-CD matrix is expected to show more potential in further applications in glycosylation analysis.

A reversed-phase depletion coupled with HILIC glycopeptide enrichment strategy by using Click OEG-CD matrix was developed in the work

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Acknowledgements

This work was financially supported by the National Natural Science Funds for Distinguished Young Scholar (20825518), the National Science Foundation of China (20805046), and the Knowledge Innovation Program of the Chinese Academy of Sciences (KSCX2-YW-R-170).

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Authors and Affiliations

  1. Key Lab of Separation Science for Analytical Chemistry, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, 116023, China

    Yanyan Zhao, Long Yu, Zhimou Guo, Xiuling Li & Xinmiao Liang

  2. College of Pharmacy, Dalian Medical University, 9 Western Lvshun South Road, Dalian, 116044, China

    Yanyan Zhao

Authors
  1. Yanyan Zhao
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  2. Long Yu
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  3. Zhimou Guo
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  4. Xiuling Li
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  5. Xinmiao Liang
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Corresponding authors

Correspondence to Xiuling Li or Xinmiao Liang.

Additional information

Yanyan Zhao and Long Yu have contributed equally to this work.

Published in the special issue Analytical and Bioanalytical Science in China with Guest Editors Lihua Zhang, Qiankun Zhuang and Yukui Zhang.

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Zhao, Y., Yu, L., Guo, Z. et al. Reversed-phase depletion coupled with hydrophilic affinity enrichment for the selective isolation of N-linked glycopeptides by using Click OEG-CD matrix. Anal Bioanal Chem 399, 3359–3365 (2011). https://doi.org/10.1007/s00216-011-4652-9

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  • DOI: https://doi.org/10.1007/s00216-011-4652-9

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