Comparison of HPLC/ESI-FTICR MS versus MALDI-TOF/TOF MS for glycopeptide analysis of a highly glycosylated HIV envelope glycoprotein

  • Janet Irungu
  • Eden P. Go
  • Ying Zhang
  • Dilusha S. Dalpathado
  • Hua-Xin Liao
  • Barton F. Haynes
  • Heather Desaire
Articles

Abstract

Defining the structures and locations of the glycans attached on secreted proteins and virus envelope proteins is important in understanding how glycosylation affects their biological properties. Glycopeptide mass spectrometry (MS)-based analysis is a very powerful, emerging approach to characterize glycoproteins, in which glycosylation sites and the corresponding glycan structures are elucidated in a single MS experiment. However, to date there is not a consensus regarding which mass spectrometric platform provides the best glycosylation coverage information. Herein, we employ two of the most widely used MS approaches, online high performance liquid chromatography-electrospray ionization mass spectrometry (HPLC/ESI-MS) and offline HPLC followed by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS), to determine which of the two approaches provides the best glycosylation coverage information of a complex glycoprotein, the group M consensus HIV-1 envelope, CON-S gp140ΔCFI, which has 31 potential glycosylation sites. Our results highlight differences in the informational content obtained between the two methods such as the overall number of glycosylation sites detected, the numbers of N-linked glycans present at each site, and the type of confirmatory information obtained about the glycopeptide using MS/MS experiments. The two approaches are quite complementary, both in their coverage of glycopeptides and in the information they provide in MS/MS experiments. The information in this study contributes to the field of mass spectrometry by demonstrating the strengths and limitations of two widely used MS platforms in glycoprotein analysis.

Supplementary material

13361_2011_190801209_MOESM1_ESM.doc (846 kb)
Supplementary material, approximately 866 KB.

References

  1. 1.
    Apweiler, R.; Hermjakob, H.; Sharon, N. On the Frequency of Protein Glycosylation, as Deduced from Analysis of the SWISS-PROT Database. Biochim. Biophys. Acta 1999, 1473, 4–8.CrossRefGoogle Scholar
  2. 2.
    Moens, S.; Vanderleyden, J. Glycoproteins in Prokaryotes. Arch. Microbiol. 1997, 168, 169–175.CrossRefGoogle Scholar
  3. 3.
    Morelle, W.; Michalski, J.-C. Glycomics and Mass Spectrometry. Curr. Pharm. Des 2005, 11, 2615–2645.CrossRefGoogle Scholar
  4. 4.
    Baenziger, J. U. The Role of Glycosylation in Protein Recognition. Am. J. Pathol. 1985, 121, 382–391.Google Scholar
  5. 5.
    Rudd, P. M.; Wormald, M. R.; Dwek, R. A. Sugar-Mediated Ligand-Receptor Interactions in the Immune System. Trends Biotechnol. 2004, 22, 524–530.CrossRefGoogle Scholar
  6. 6.
    Wang, L.-X. Toward Oligosaccharide- and Glycopeptide-Based HIV Vaccines. Curr. Opin. Drug Discov. Devel. 2006, 9, 194–206.Google Scholar
  7. 7.
    Geyer, H.; Holschbach, C.; Hunsmann, G.; Schneider, J. Carbohydrates of Human Immunodeficiency Virus: Structures of Oligosaccharides Linked to the Envelope Glycoprotein 120. J. Biol. Chem. 1988, 263, 11760–11767.Google Scholar
  8. 8.
    Koch, M.; Pancera, M.; Kwong, P. D.; Kolchinsky, P.; Grundner, C.; Wang, L.; Hendrickson, W. A.; Sodroski, J.; Wyatt, R. Structure-Based, Targeted Deglycosylation of HIV-1 gp120 and Effects on Neutralization Sensitivity and Antibody Recognition. Virology 2003, 313, 387–400.CrossRefGoogle Scholar
  9. 9.
    Leonard, C. K.; Spellman, M. W.; Riddle, L.; Harris, R. J.; Thomas, J. N.; Gregory, T. J. Assignment of Intrachain Disulfide Bonds and Characterization of Potential Glycosylation Sites of the Type 1 Recombinant Human Immunodeficiency Virus Envelope Glycoprotein (gp120) Expressed in Chinese Hamster Ovary Cells. J. Biol. Chem. 1990, 265, 10373–10382.Google Scholar
  10. 10.
    Mizuochi, T.; Matthews, T. J.; Kato, M.; Hamako, J.; Titani, K.; Solomon, J.; Feizi, T. Diversity of Oligosaccharide Structures on the Envelope Glycoprotein gp120 of Human Immunodeficiency Virus 1 from the Lymphoblastoid Cell Line H9: Presence of Complex-Type Oligosaccharides with Bisecting N- Acetylglucosamine Residues. J. Biol. Chem. 1990, 265, 8519–8524.Google Scholar
  11. 11.
    Back, N. K.; Smit, L.; De Jong, J. J.; Keulen, W.; Schutten, M.; Goudsmit, J.; Tersmette, M. An N-Glycan Within the Human Immunodeficiency Virus Type 1 gp120 V3 Loop Affects Virus Neutralization. Virology 1994, 199, 431–438.CrossRefGoogle Scholar
  12. 12.
    Cheng-Mayer, C.; Brown, A.; Harouse, J.; Luciw, P. A.; Mayer, A. J. Selection for Neutralization Resistance of the Simian/Human Immunodeficiency Virus SHIVSF33A Variant in Vivo by Virtue of Sequence Changes in the Extracellular Envelope Glycoprotein that Modify N-Linked Glycosylation. J. Virol. 1999, 73, 5294–5300.Google Scholar
  13. 13.
    Kwong, P. D.; Doyle, M. L.; Casper, D. J.; Cicala, C.; Leavitt, S. A.; Majeed, S.; Steenbeke, T. D.; Venturi, M.; Chaiken, I.; Fung, M.; Katinger, H.; Parren, P. W. I. H.; Robinson, J.; Van Ryk, D.; Wang, L.; Burton, D. R.; Freire, E.; Wyatt, R.; Sodroski, J.; Hendrickson, W. A.; Arthos, J. HIV-1 Evades Antibody-Mediated Neutralization Through Conformational Masking of Receptor-Binding Sites. Nature 2002, 420, 678–682.CrossRefGoogle Scholar
  14. 14.
    Reitter, J. N.; Means, R. E.; Desrosiers, R. C. A Role for Carbohydrates in Immune Evasion in AIDS. Nat. Med. 1998, 4, 679–684.CrossRefGoogle Scholar
  15. 15.
    Wyatt, R.; Kwong, P. D.; Desjardins, E.; Sweet, R. G.; Robinson, J.; Hendrickson, W. A.; Sodroski, J. G. The Antigenic Structure of the HIV gp120 Envelope Glycoprotein. Nature 1998, 393, 705–711.CrossRefGoogle Scholar
  16. 16.
    Zhu, X.; Borchers, C.; Bienstock, R. J.; Tomer, K. B. Mass Spectrometric Characterization of the Glycosylation Pattern of HIV-gp120 Expressed in CHO Cells. Biochemistry 2000, 39, 11194–11204.CrossRefGoogle Scholar
  17. 17.
    Yeh, J.-C.; Seals, J. R.; Murphy, C. I.; van Halbeek, H.; Cummings, R. D. Site-Specific N-Glycosylation and Oligosaccharide Structures of Recombinant: HIV-1 gp120 Derived from a Baculovirus Expression system. Biochemistry 1993, 32, 11087–11099.CrossRefGoogle Scholar
  18. 18.
    Morelle, W.; Canis, K.; Chirat, F.; Faid, V.; Michalski, J.-C. The Use of Mass Spectrometry for the Proteomic Analysis of Glycosylation. Proteomics 2006, 6, 3993–4015.CrossRefGoogle Scholar
  19. 19.
    Wada, Y.; Azadi, P.; Costello, C. E.; Dell, A.; Dwek, R. A.; Geyer, H.; Geyer, R.; Kakehi, K.; Karlsson, N. G.; Kato, K.; Kawasaki, N.; Khoo, K.-H.; Kim, S.; Kondo, A.; Lattova, E.; Mechref, Y.; Miyoshi, E.; Nakamura, K.; Narimatsu, H.; Novotny, M. V.; Packer, N. H.; Perreault, H.; Peter-Katalinic, J.; Pohlentz, G.; Reinhold, V. N.; Rudd, P. M.; Suzuki, A.; Taniguchi, N. Comparison of the Methods for Profiling Glycoprotein Glycans—HUPO Human Disease Glycomics/Proteome Initiative Multi-Institutional Study. Glycobiology 2007, 17, 411–422.CrossRefGoogle Scholar
  20. 20.
    Budnik, B. A.; Lee, R. S.; Steen, J. A. J. Global Methods for Protein Glycosylation Analysis by Mass Spectrometry. Biochim. Biophys. Acta 2006, 1764, 1870–1880.CrossRefGoogle Scholar
  21. 21.
    Mechref, Y.; Novotny, M. V. Structural Investigations of Glycoconjugates at High Sensitivity. Chem. Rev. 2002, 102, 321–369.CrossRefGoogle Scholar
  22. 22.
    Wuhrer, M.; Koeleman, C. A. M.; Hokke, C. H.; Deelder, A. M. Protein Glycosylation Analyzed by Normal-Phase Nano-Liquid Chromatography Mass Spectrometry of Glycopeptides. Anal. Chem. 2005, 77, 886–894.CrossRefGoogle Scholar
  23. 23.
    Carr, S. A.; Huddleston, M. J.; Bean, M. F. Selective Identification and Differentiation of N- and O-Linked Oligosaccharides in Glycoproteins by Liquid Chromatography-Mass Spectrometry. Protein Sci. 1993, 2, 183–196.CrossRefGoogle Scholar
  24. 24.
    Stephens, E.; Maslen, S. L.; Green, L. G.; Williams, D. H. Fragmentation Characteristics of Neutral N-Linked Glycans Using a MALDI-TOF/TOF Tandem Mass Spectrometer. Anal. Chem. 2004, 76, 2343–2354.CrossRefGoogle Scholar
  25. 25.
    Burlingame, A. L. Characterization of Protein Glycosylation by Mass Spectrometry. Curr. Opin. Biotechnol. 1996, 7, 4–10.CrossRefGoogle Scholar
  26. 26.
    Kurogochi, M.; Nishimura, S. I. Structural Characterization of N-Glycopeptides by Matrix-Dependent Selective Fragmentation of MALDI-TOF/TOF Tandem Mass Spectrometry. Anal. Chem. 2004, 76, 6097–6101.CrossRefGoogle Scholar
  27. 27.
    Wuhrer, M.; Catalina, M. I.; Deelder, A. M.; Hokke, C. H. Glycoproteomics Based on Tandem Mass Spectrometry of Glycopeptides. J. Chromatogr. B Analyt. Technol. Biomed. Life Sci. 2007, 849, 115–128.CrossRefGoogle Scholar
  28. 28.
    Go, E. P.; Irungu, J.; Zhang, Y.; Dalpathado, D. S.; Liao, H.-X.; Sutherland, L. L.; Alam, S. M.; Haynes, B. F.; Desaire, H. Glycosylation Site-Specific Analysis of HIV Envelope Proteins (JR-FL and CON-S) Reveals Major Differences in Glycosylation Site Occupancy, Glycoform Profiles, and Antigenic Epitopes’ Accessibility. J. Proteome 2008, 7(4), 1660–1674.CrossRefGoogle Scholar
  29. 29.
    Liao, H.-X.; Sutherland, L. L.; Xia, S.-M.; Brock, M. E.; Scearce, R. M.; Vanleeuwen, S.; Alam, S. M.; McAdams, M.; Weaver, E. A.; Camacho, Z. T.; Ma, B.-J.; Li, Y.; Decker, J. M.; Nabel, G. J.; Montefiori, D. C.; Hahn, B. H.; Korber, B. T.; Gao, F.; Haynes, B. F. A Group M Consensus Envelope Glycoprotein Induces Antibodies that Neutralize Subsets of Subtype B and C HIV-1 Primary Viruses. Virology 2006, 353, 268–282.CrossRefGoogle Scholar
  30. 30.
    Cutalo, J. M.; Deterding, L. J.; Tomer, K. B. Characterization of Glycopeptides from HIV-ISF2 gp120 by Liquid Chromatography Mass Spectrometry. J. Am. Soc. Mass Spectrom. 2004, 15, 1545–1555.CrossRefGoogle Scholar
  31. 31.
    Go, E. P.; Rebecchi, K. R.; Dalpathado, D. S.; Bandu, M. L.; Zhang, Y.; Desaire, H. GlycoPep DB: A Tool for Glycopeptide Analysis Using a “Smart Search”. Anal. Chem. 2007, 79, 1708–1713.CrossRefGoogle Scholar
  32. 32.
    Irungu, J.; Go, E. P.; Dalpathado, D. S.; Desaire, H. Simplification of Mass Spectral Analysis of Acidic Glycopeptides Using GlycoPep ID. Anal. Chem. 2007, 79, 3065–3074.CrossRefGoogle Scholar
  33. 33.
    Wuhrer, M.; Hokke, C. H.; Deelder, A. M. Glycopeptide Analysis by Matrix-Assisted Laser Desorption/Ionization Tandem Time-of-Flight Mass Spectrometry Reveals Novel Features of Horseradish Peroxidase Glycosylation. Rapid Commun. Mass Spectrom 2004, 18, 1741–1748.CrossRefGoogle Scholar
  34. 34.
    Irungu, J.; Dalpathado, D. S.; Go, E. P.; Jiang, H.; Ha, H.-V.; Bousfield, G. R.; Desaire, H. Method for Characterizing Sulfated Glycoproteins in a Glycosylation Site-Specific Fashion, Using Ion Pairing and Tandem Mass Spectrometry. Anal. Chem. 2006, 78, 1181–1190.CrossRefGoogle Scholar
  35. 35.
    Dalpathado, D. S.; Desaire, H. Glycopeptide Analysis by Mass Spectrometry. Analyst 2008, 133, 731–738.CrossRefGoogle Scholar
  36. 36.
    Carr, S. A.; Roberts, G. D. Carbohydrate Mapping by Mass Spectrometry: A Novel Method for Identifying Attachment Sites of Asn-Linked Sugars in Glycoproteins. Anal. Biochem. 1986, 157, 396–406.CrossRefGoogle Scholar
  37. 37.
    Brancia, F. L.; Oliver, S. G.; Gaskell, S. J. Improved Matrix-Assisted Laser Desorption/Ionization Mass Spectrometric Analysis of Tryptic Hydrolysates of Proteins Following Guanidination of Lysine-Containing Peptides. Rapid Commun. Mass Spectrom. 2000, 14, 2070–2073.CrossRefGoogle Scholar

Copyright information

© American Society for Mass Spectrometry 2008

Authors and Affiliations

  • Janet Irungu
    • 1
  • Eden P. Go
    • 1
  • Ying Zhang
    • 1
  • Dilusha S. Dalpathado
    • 1
  • Hua-Xin Liao
    • 2
  • Barton F. Haynes
    • 2
  • Heather Desaire
    • 1
  1. 1.Department of ChemistryUniversity of KansasLawrenceUSA
  2. 2.Duke University Medical CenterDurhamUSA

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