Advertisement

Analysis of Glycopeptides Using Porous Graphite Chromatography and LTQ Orbitrap XL ETD Hybrid MS

  • Terry Zhang
  • Rosa Viner
  • Zhiqi Hao
  • Vlad ZabrouskovEmail author
Chapter

Abstract

In this study, several glycoproteins: bovine α1-acid glycoprotein, fetuin, and human α1-acid glycoprotein, were analyzed using nanoLC-MS/MS. The performance of three different stationary phases – C8, C18, and porous graphite – was systematically evaluated and optimized for glycopeptide separation. A porous graphite column was chosen as the optimum stationary phase for glycoprotein analysis. An LTQ Orbitrap XL hybrid mass spectrometer equipped with electron transfer dissociation (ETD) was used to acquire, within a single analysis, high-resolution, high-mass-accuracy full MS spectra, collision-induced dissociation (CID) MS/MS spectra for glycan composition analysis, and ETD MS/MS spectra for peptide structure information. As a result, characterization of glycopeptides was achieved within a single LC-MS/MS run.

Keywords

ETD Glycoprotein N-glycosylation LTQ Orbitrap XL ETD Porous graphite 

Notes

Acknowledgement

We thank David Fisher at Thermo Fisher Scientific for editing and proof reading of this manuscript.

References

  1. Alley Jr., W.R., Merchref, Y., and Novotny, M.V. (2007). Using Graphitized Carbon for Glycopeptides Separations Prior to Mass Spectral Detection. Proceedings of the 55th ASMS Conference (Indianapolis, IN).Google Scholar
  2. Alley Jr., W.R., Merchref, Y., and Novotny, M.V. (2009). Characterization of glycopeptides by combining collision-induced dissociation and electron-transfer dissociation mass spectrometry data. Rapid Commun Mass Spectrom 23, 161–170.CrossRefGoogle Scholar
  3. Kaji, H, and Isobe, T. (2008). Liquid chromatography/mass spectrometry (LC-MS)-based glycoproteomics technologies for cancer biomarker discovery. Clin Proteomics 4, 14–24.CrossRefGoogle Scholar
  4. McAlister, G.C., Phanstiel, D., Good D.M., Berggren, W.T., and Coon, J.J. (2007). Implementation of electron-transfer dissociation on a hybrid linear ion trap-orbitrap mass spectrometer. Mol Cell Proteomics 6, 1942–1951.CrossRefGoogle Scholar
  5. Medzihradszky, K.F., Guan, S., Maltby, D.A., and Burlingame, A.L. (2007). Sulfopeptide fragmentation in electron-capture and electron-transfer dissociation. J Am Soc Mass Spectro 18(9), 1617–1624CrossRefGoogle Scholar
  6. Morelle, W., Canis, K., Chirat, F., Faid, V., and Michalski, J-C. (2006). The use of mass spectrometry for the proteomic analysis of glycosylation. Proteomics 6, 3993–3915.Google Scholar
  7. Newton, K.A., Amunugama, R., and McLuckey, S.A. (2005). Gas-phase ion/ion reactions of multiply protonated polypeptides with metal containing anions. J Phys Chem A 109(16), 3608–3616.CrossRefGoogle Scholar
  8. Peterman, S.M., and Mulholland, J.J. (2006). A novel approach for identification and characterization of glycoproteins using a hybrid linear ion trap/FT-ICR mass spectrometer. J Am Soc Mass Spectrom 17(2), 68–79.CrossRefGoogle Scholar
  9. Snovida, S.I., Chen, V.C., Krokhin, O., and Perreault, H. (2006). Isolation and identification of sialylated glycopeptides from bovine α1-acid glycoprotein by off-line capillary electrophoresis MALDI-TOF mass spectrometry. Anal Chem 78, 6556–635CrossRefGoogle Scholar
  10. Syka J.E., Coon J.J., Schroder M.J., Shabanowitz J, and Hunt D.F. (2004). Peptide and protein sequence analysis by electron transfer dissociation mass spectrometry. Proc Natl Acad Sci USA 101, 9528–9533.CrossRefGoogle Scholar
  11. Treuheit, M.J., Costello, C.E., and Halsall, H.B. (1992). Analysis of the five glycosylation sites of human α1-acid glycoprotein. Biochem J 283, 105–112.Google Scholar

Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • Terry Zhang
    • 1
  • Rosa Viner
    • 1
  • Zhiqi Hao
    • 1
  • Vlad Zabrouskov
    • 1
    Email author
  1. 1.Thermo Fisher ScientificSan JoseUSA

Personalised recommendations