Glycan size and attachment site location affect electron transfer dissociation (ETD) fragmentation and automated glycopeptide identification

  • Kathirvel Alagesan
  • Hannes Hinneburg
  • Peter H. Seeberger
  • Daniel Varón Silva
  • Daniel KolarichEmail author
Short Communication




Glycopeptide Glycoproteomics Electron transfer dissociation ETD 



We thank the Beilstein-Institut for supporting KA with a PhD scholarship and the Max Planck Society for financial support. DK is the recipient of an Australian Research Council Future Fellowship (project number FT160100344) funded by the Australian Government.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflicts of interest.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Supplementary material

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ESM 1 (PDF 383 kb)
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  1. 1.
    Alley Jr., W.R., Mechref, Y., Novotny, M.V.: Characterization of glycopeptides by combining collision-induced dissociation and electron-transfer dissociation mass spectrometry data. Rapid Commun. Mass Spectrom. 23(1), 161–170 (2009)CrossRefGoogle Scholar
  2. 2.
    Mechref, Y., Use of CID/ETD mass spectrometry to analyze glycopeptides. Curr Protoc Protein Sci, 2012. Chapter 12: p. Unit 12 11 1–11Google Scholar
  3. 3.
    Segu, Z.M., Mechref, Y.: Characterizing protein glycosylation sites through higher-energy C-trap dissociation. Rapid Commun. Mass Spectrom. 24(9), 1217–1225 (2010)CrossRefGoogle Scholar
  4. 4.
    Palmisano, G., Larsen, M.R., Packer, N.H., Thaysen-Andersen, M.: Structural analysis of glycoprotein sialylation -part II: LC-MS based detection. RSC Adv. 3(45), 22706–22726 (2013)CrossRefGoogle Scholar
  5. 5.
    Zhurov, K.O., Fornelli, L., Wodrich, M.D., Laskay, Ü.A., Tsybin, Y.O.: Principles of electron capture and transfer dissociation mass spectrometry applied to peptide and protein structure analysis. Chem. Soc. Rev. 42(12), 5014–5030 (2013)CrossRefGoogle Scholar
  6. 6.
    Syrstad, E.A., Turecek, F.: Toward a general mechanism of electron capture dissociation. J. Am. Soc. Mass Spectrom. 16(2), 208–224 (2005)CrossRefGoogle Scholar
  7. 7.
    Wuhrer, M., Catalina, M.I., Deelder, A.M., Hokke, C.H.: Glycoproteomics based on tandem mass spectrometry of glycopeptides. J. Chromatogr. B Anal. Technol. Biomed. Life Sci. 849(1–2), 115–128 (2007)CrossRefGoogle Scholar
  8. 8.
    Hinneburg, H., Stavenhagen, K., Schweiger-Hufnagel, U., Pengelley, S., Jabs, W., Seeberger, P.H., Silva, D.V., Wuhrer, M., Kolarich, D.: The art of destruction: optimizing collision energies in Quadrupole-Time of Flight (Q-TOF) instruments for glycopeptide-based Glycoproteomics. J. Am. Soc. Mass Spectrom. 27(3), 507–519 (2016)CrossRefGoogle Scholar
  9. 9.
    Yamamoto, N., Ohmori, Y., Sakakibara, T., Sasaki, K., Juneja, L.R., Kajihara, Y.: Solid-phase synthesis of sialylglycopeptides through selective esterification of the sialic acid residues of an Asn-linked complex-type sialyloligosaccharide. Angew. Chem. Int. Ed. Engl. 42(22), 2537–2540 (2003)CrossRefGoogle Scholar
  10. 10.
    Alagesan, K., Khilji, S.K., Kolarich, D.: It is all about the solvent: on the importance of the mobile phase for ZIC-HILIC glycopeptide enrichment. Anal. Bioanal. Chem. 409(2), 529–538 (2017)CrossRefGoogle Scholar
  11. 11.
    Stavenhagen, K., Hinneburg, H., Thaysen-Andersen, M., Hartmann, L., Silva, D.V., Fuchser, J., Kaspar, S., Rapp, E., Seeberger, P.H., Kolarich, D.: Quantitative mapping of glycoprotein micro-heterogeneity and macro-heterogeneity: an evaluation of mass spectrometry signal strengths using synthetic peptides and glycopeptides. J. Mass Spectrom. 48(6), 627–639 (2013)CrossRefGoogle Scholar
  12. 12.
    Hinneburg, H., Hofmann, J., Struwe, W.B., Thader, A., Altmann, F., Varón Silva, D., Seeberger, P.H., Pagel, K., Kolarich, D.: Distinguishing N-acetylneuraminic acid linkage isomers on glycopeptides by ion mobility-mass spectrometry. Chem. Commun. (Camb.). 52(23), 4381–4384 (2016)CrossRefGoogle Scholar
  13. 13.
    Piontek, C., Ring, P., Harjes, O., Heinlein, C., Mezzato, S., Lombana, N., Pöhner, C., Püttner, M., Varón Silva, D., Martin, A., Schmid, F.X., Unverzagt, C.: Semisynthesis of a homogeneous glycoprotein enzyme: ribonuclease C: part 1. Angew. Chem. Int. Ed. Engl. 48(11), 1936–1940 (2009)CrossRefGoogle Scholar
  14. 14.
    Gil, G.C., Velander, W.H., Van Cott, K.E.: N-glycosylation microheterogeneity and site occupancy of an Asn-X-Cys sequon in plasma-derived and recombinant protein C. Proteomics. 9(9), 2555–2567 (2009)CrossRefGoogle Scholar
  15. 15.
    Kolarich, D., Rapp, E., Struwe, W.B., Haslam, S.M., Zaia, J., McBride, R., Agravat, S., Campbell, M.P., Kato, M., Ranzinger, R., Kettner, C., York, W.S.: The minimum information required for a glycomics experiment (MIRAGE) project: improving the standards for reporting mass-spectrometry-based glycoanalytic data. Mol. Cell. Proteomics. 12(4), 991–995 (2013)CrossRefGoogle Scholar
  16. 16.
    Taylor, C.F., Paton, N.W., Lilley, K.S., Binz, P.A., Julian, R.K., Jones, A.R., Zhu, W., Apweiler, R., Aebersold, R., Deutsch, E.W., Dunn, M.J., Heck, A.J.R., Leitner, A., Macht, M., Mann, M., Martens, L., Neubert, T.A., Patterson, S.D., Ping, P., Seymour, S.L., Souda, P., Tsugita, A., Vandekerckhove, J., Vondriska, T.M., Whitelegge, J.P., Wilkins, M.R., Xenarios, I., Yates, J.R., Hermjakob, H.: The minimum information about a proteomics experiment (MIAPE). Nat. Biotechnol. 25(8), 887–893 (2007)CrossRefGoogle Scholar
  17. 17.
    Liu, J., McLuckey, S.A.: Electron transfer dissociation: effects of cation charge state on product partitioning in ion/ion electron transfer to multiply protonated polypeptides. Int. J. Mass Spectrom. 330-332, 174–181 (2012)CrossRefGoogle Scholar
  18. 18.
    Lin, C.W., Haeuptle, M.A., Aebi, M.: Supercharging reagent for enhanced liquid chromatographic separation and charging of sialylated and high-molecular-weight glycopeptides for NanoHPLC-ESI-MS/MS analysis. Anal. Chem. 88(17), 8484–8494 (2016)CrossRefGoogle Scholar
  19. 19.
    Windwarder, M., Yelland, T., Djordjevic, S., Altmann, F.: Detailed characterization of the O-linked glycosylation of the neuropilin-1 c/MAM-domain. Glycoconj. J. 33(3), 387–397 (2016)CrossRefGoogle Scholar
  20. 20.
    Baker, P.R., Medzihradszky, K.F., Chalkley, R.J.: Improving software performance for peptide electron transfer dissociation data analysis by implementation of charge state- and sequence-dependent scoring. Mol. Cell. Proteomics. 9(9), 1795–1803 (2010)CrossRefGoogle Scholar
  21. 21.
    Zheng, K., Yarmarkovich, M., Bantog, C., Bayer, R., Patapoff, T.W.: Influence of glycosylation pattern on the molecular properties of monoclonal antibodies. MAbs. 6(3), 649–658 (2014)CrossRefGoogle Scholar
  22. 22.
    Xia, Y., Gunawardena, H.P., Erickson, D.E., McLuckey, S.A.: Effects of cation charge-site identity and position on electron-transfer dissociation of polypeptide cations. J. Am. Chem. Soc. 129(40), 12232–12243 (2007)CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Institute for GlycomicsGriffith UniversitySouthportAustralia
  2. 2.Department of Biomolecular SystemsMax Planck Institute of Colloids and InterfacesPotsdamGermany
  3. 3.Institute of Chemistry and BiochemistryFreie Universität BerlinBerlinGermany
  4. 4.Department of Molecular Science, Faculty of Science and EngineeringMacquarie UniversityNorth RydeAustralia
  5. 5.ARC Centre for Nanoscale BioPhotonicsAdelaideAustralia

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