Structural characterization of the GM1 ganglioside by infrared multiphoton dissociation, electron capture dissociation, and electron detachment dissociation electrospray ionization FT-ICR MS/MS

  • Melinda A. McFarland
  • Alan G. Marshall
  • Christopher L. Hendrickson
  • Carol L. Nilsson
  • Pam Fredman
  • Jan-Eric Månsson


Gangliosides play important biological roles and structural characterization of both the carbohydrate and the lipid moieties is important. The FT-ICR MS/MS techniques of electron capture dissociation (ECD), electron detachment dissociation (EDD), and infrared multiphoton dissociation (IRMPD) provide extensive fragmentation of the protonated and deprotonated GM1 ganglioside. ECD provides extensive structural information, including identification of both halves of the ceramide and cleavage of the acetyl moiety of the N-acetylated sugars. IRMPD provides similar glycan fragmentation but no cleavage of the acetyl moiety. Cleavage between the fatty acid and the long-chain base of the ceramide moiety is seen in negative-ion IRMPD but not in positive-ion IRMPD of GM1. Furthermore, this extent of fragmentation requires a range of laser powers, whereas all information is available from a single ECD experiment. However, stepwise fragmentation by IRMPD may be used to map the relative labilities for a series of cleavages. EDD provides the alternative of electron-induced fragmentation for negative ions with extensive fragmentation, but suffers from low efficiency as well as complication of data analysis by frequent loss of hydrogen atoms. We also show that analysis of MS/MS data for glycolipids is greatly simplified by classification of product ion masses to specific regions of the ganglioside based solely on mass defect graphical analysis.


  1. 1.
    Svennerholm, L. Designation and Schematic Structure of Gangliosides and Allied Glycosphingolipids Biological Function of Gangliosides; Vol. CI; Elsevier Science: Amsterdam, 1994; pp R11-R14.Google Scholar
  2. 2.
    Karlsson, K. A. Animal Glycosphingolipids as Membrane Attachment Sites for Bacteria. Annu. Rev. Biochem. 1989, 58, 309–350.CrossRefGoogle Scholar
  3. 3.
    Hakomori, S. Traveling for the Glycosphingolipid Path. Glycoconj. J. 2000, 17, 627–647.CrossRefGoogle Scholar
  4. 4.
    Futerman, A. H.; Hannun, Y. A. The Complex Life of Simple Sphingolipids. EMBO. J. 2004, 8, 777–782.Google Scholar
  5. 5.
    Svennerholm, L.; Fredman, P. A. Procedure for the Quantitative Isolation of Brain Gangliosides. Biochim. Biophys. Acta 1980, 617, 97–109.Google Scholar
  6. 6.
    Mansson, J. E.; Holmgren, J.; Li, Y. T.; Vanier, M. T.; Svennerholm, L. Chemical and Immunological Characterization of the Major Glucosamine-Containing Gangliosides of Human Tissues. Med. Biol. 1974, 52, 240–243.Google Scholar
  7. 7.
    Kannagi, R.; Nudelman, E.; Hakomori, S. I. Possible Role of Ceramide in Defining Structure and Function of Membrane Glycolipids. Proc. Nat. Acad. Sci. U.S.A. 1982, 79, 3470–3474.CrossRefGoogle Scholar
  8. 8.
    Mansson, J. E.; Svennerholm, L. The Use of Galactosylceramides with Uniform Fatty Acids as Substrates in the Diagnosis and Carrier Detection of Krabbe Disease. Clin. Chim. Acta 1982, 126, 127–133.CrossRefGoogle Scholar
  9. 9.
    Lingwood, C. A. Aglycone Modulation of Glycolipid Receptor Function. Glycoconj. J. 1996, 13, 495–503.CrossRefGoogle Scholar
  10. 10.
    Egge, H.; Peterkatalinic, J.; Reuter, G.; Schauer, R.; Ghidoni, R.; Sonnino, S.; Tettamanti, G. Analysis of Gangliosides Using Fast Atom Bombardment Mass-Spectrometry. Chem. Phys. Lipids 1985, 37, 127–141.CrossRefGoogle Scholar
  11. 11.
    Ladisch, S.; Sweeley, C. C.; Becker, H.; Gage, D. Aberrant Fatty Acyl α-Hydroxylation in Human Neuro-Blastoma Tumor Gangliosides. J. Biol. Chem. 1989, 264, 12097–12105.Google Scholar
  12. 12.
    Domon, B.; Costello, C. E. Structure Elucidation of Glycosphingolipids and Gangliosides Using High-Performance Tandem Mass-Spectrometry. Biochemistry 1988, 27, 1534–1543.CrossRefGoogle Scholar
  13. 13.
    Juhasz, P.; Costello, C. E. Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass-Spectrometry of Underivatized and Permethylated Gangliosides. J. Am. Soc. Mass Spectrom 1992, 3, 785–796.CrossRefGoogle Scholar
  14. 14.
    Yohe, H. C.; Wallace, P. K.; Berenson, C. S.; Ye, S.; Reinhold, B. B.; Reinhold, V. N. The Major Gangliosides of Human Peripheral Blood Monocytes/Macrophages: Absence of Ganglio Series Structures. Glycobiology 2001, 11, 831–841.CrossRefGoogle Scholar
  15. 15.
    Metelmann, W.; Peter-Katalinic, J.; Muthing, J. Gangliosides from Human Granulocytes: A Nano-ESI QTOF Mass Spectrometry Fucosylation Study of Low Abundance Species in Complex Mixtures. J. Am. Soc. Mass Spectrom. 2001, 12, 964–973.CrossRefGoogle Scholar
  16. 16.
    Metelmann, W.; Vukelic, Z.; Peter-Katalinic, J. Nano-Electrospray Ionization Time-of-flight Mass Spectrometry of Gangliosides from Human Brain Tissue. J. Mass Spectrom. 2001, 36, 21–29.CrossRefGoogle Scholar
  17. 17.
    Isaac, G.; Bylund, D.; Mansson, J. E.; Markides, K. E.; Bergquist, J. Analysis of Phosphatidylcholine and Sphingomyelin Molecular Species from Brain Extracts using Capillary Liquid Chromatography Electrospray Ionization Mass Spectrometry. J. Neurosci. Methods 2003, 128, 111–119.CrossRefGoogle Scholar
  18. 18.
    Marshall, A. G.; Hendrickson, C. L.; Jackson, G. S. Fourier Transform Ion Cyclotron Resonance Mass Spectrometry: A Primer. Mass Spectrom. Rev. 1998, 17, 1–35.CrossRefGoogle Scholar
  19. 19.
    Marshall, A. G. Milestones in Fourier Transform Ion Cyclotron Resonance Mass Spectrometry Technique Development. Int. J. Mass Spectrom 2000, 200, 331–356.CrossRefGoogle Scholar
  20. 20.
    Gauthier, J. W.; Trautman, T. R.; Jacobson, D. B. Sustained Off-Resonance Irradiation for CAD Involving FTMS. CAD Technique that Emulates Infrared Multiphoton Dissociation. Anal. Chim. Acta 1991, 246, 211–225.CrossRefGoogle Scholar
  21. 21.
    McLuckey, S. A. Principles of Collisional Activation in Analytical Mass-Spectrometry. J. Am. Soc. Mass Spectrom. 1992, 3, 599–614.CrossRefGoogle Scholar
  22. 22.
    Senko, M. W.; Speir, J. P.; McLafferty, F. W. Collisional Activition of Large Multiply-Charged Ions Using Fourier Transform Mass Spectrometry. Anal. Chem. 1994, 66, 2801–2808.CrossRefGoogle Scholar
  23. 23.
    Woodlin, R. L.; Bomse, D. S.; Beauchamp, J. L. Multiphoton Dissociation of Molecules with Low Power Continuous Wave Infrared Laser Radiation. J. Am. Chem. Soc. 1978, 100, 3248–3250.CrossRefGoogle Scholar
  24. 24.
    Little, D. P.; Speir, J. P.; Senko, M. W.; O’Connor, P. B.; McLafferty, F. W. Infrared Multiphoton Dissociation of Large Multiply-Charged Ions For Biomolecule Sequencing. Anal. Chem. 1994, 66, 2809–2815.CrossRefGoogle Scholar
  25. 25.
    Zubarev, R. A.; Kelleher, N. L.; McLafferty, F. W. Electron Capture Dissociation of Multiply Charged Protein Cations. A Nonergodic Process. J. Am. Chem. Soc. 1998, 120, 3265–3266.CrossRefGoogle Scholar
  26. 26.
    Zubarev, R. A.; Horn, D. M.; Fridriksson, E. K.; Kelleher, N. L.; Kruger, N. A.; Lewis, M. A.; Carpenter, B. K.; McLafferty, F. W. Electron Capture Dissociation for Structural Characterization of Multiply Charged Protein Cations. Anal. Chem. 2000, 72, 563–573.CrossRefGoogle Scholar
  27. 27.
    Budnik, B. A.; Haselmann, K. F.; Zubarev, R. A. Electron Detachment Dissociation of Peptide Di-Anions: An Electron-Hole Recombination Phenomenon. Chem. Phys. Lett. 2001, 342, 299–302.CrossRefGoogle Scholar
  28. 28.
    Gaucher, S. P.; Cancilla, M. T.; Phillips, N. J.; Gibson, B. W.; Leary, J. A. Mass Spectral Characterization of Lipooligosaccharides from Haemophilus influenzae 2019. Biochemistry 2000, 39, 12406–12414.CrossRefGoogle Scholar
  29. 29.
    Mirgorodskaya, E.; O’Connor, P. B.; Costello, C. E. A General Method for Precalculation of Parameters for Sustained Off-Resonance Irradiation/Collision-Induced Dissociation. J. Am. Soc. Mass Spectrom. 2002, 13, 318–324.CrossRefGoogle Scholar
  30. 30.
    Mirgorodskaya, E.; Roepstorff, P.; Zubarev, R. A. Localization of O-Glycosylation Sites in Peptides by Electron Capture Dissociation in a Fourier Transform Mass Spectrometer. Anal. Chem. 1999, 71, 4431–4436.CrossRefGoogle Scholar
  31. 31.
    Guan, Z. Identification and Localization of the Fatty Acid Modification in Ghrelin by Electron Capture Dissociation. J. Am. Soc. Mass Spectrom. 2002, 13, 1443–1447.CrossRefGoogle Scholar
  32. 32.
    Budnik, B. A.; Haselmann, K. F.; Elkin, Y. N.; Gorbach, V. I.; Zubarev, R. A. Applications of Electron-Ion Dissociation Reactions for Analysis of Polycationic Chitooligosaccharides in Fourier Transform Mass Spectrometry. Anal. Chem. 2003, 75, 5994–6001.CrossRefGoogle Scholar
  33. 33.
    Håkansson, K.; Cooper, H. J.; Emmett, M. R.; Costello, C. E.; Marshall, A. G.; Nilsson, C. L. Electron Capture Dissociation and Infrared Multiphoton Dissociation MS/MS of an N-Glycosylated Tryptic Peptide Yield Complementary Sequence Information. Anal. Chem. 2001, 73, 4530–4536.CrossRefGoogle Scholar
  34. 34.
    Mansson, J. E.; Fredman, P.; Bigner, D. D.; Molin, K.; Rosengren, B.; Friedman, H. S.; Svennerholm, L. Characterization of New Gangliosides of the Lactotetraose Series in Murine Xenografts of a Human Glioma Cell Line. FEBS Lett. 1986, 201, 109–113.CrossRefGoogle Scholar
  35. 35.
    Mansson, J. E.; Mo, H.; Egge, H.; Svennerholm, L. Trisialosyllactosylceramide (GT3) is a Ganglioside of Human Lung. FEBS Lett. 1986, 196, 259–262.CrossRefGoogle Scholar
  36. 36.
    Svennerholm, L.; Rynmark, B. M.; Vilbergson, G.; Fredman, P.; Gottfries, J.; Mansson, J. E.; Percy, A. Gangliosides in Human Brain. J. Neurochem. 1991, 56, 1763–1768.CrossRefGoogle Scholar
  37. 37.
    Senko, M. W.; Hendrickson, C. L.; PasaTolic, L.; Marto, J. A.; White, F. M.; Guan, S. H.; Marshall, A. G. Electrospray Ionization Fourier Transform Ion Cyclotron Resonance at 9.4 T. Rapid Commun. Mass Spectrom. 1996, 10, 1824–1828.CrossRefGoogle Scholar
  38. 38.
    Blakney, G.; Lam, T.; Hendrickson, C. L.; Marshall, A. G. FT-ICR MS Data Station for Automated High Speed Data-Dependent Acquisition. Proceedings of the 52nd ASMS Conference on Mass Spectrometry and Allied Topics; Nashville, TN, 2004.Google Scholar
  39. 39.
    Quinn, J. P.; Emmett, M. R.; Marshall, A. G. A Device for Fabrication of Emitters for Low-Flow Electrospray Ionization; Proceedings of the 46th ASMS Conference on Mass Spectrometry and Allied Topics, Orlando FL, May 1998; pp 1388–1388Google Scholar
  40. 40.
    Hendrickson, C. L.; Quinn, J. P.; Emmett, M. R.; Marshall, A. G. Quadrupole Mass Filtered External Accumulation for Fourier Transform Ion Cyclotron Resonance Mass Spectrometry. Proceedings of the 48th ASMS Conference on Mass Spectrometry and Allied Topics; Long Beach, CA, June 2000.Google Scholar
  41. 41.
    Hendrickson, C. L.; Quinn, J. P.; Emmett, M. R.; Marshall, A. G. Mass-Selective External Ion Accumulation for Fourier Transform Ion Cyclotron Resonance Mass Spectrometry. Proceedings of the 49th ASMS Conference on Mass Spectrometry and Allied Topics; Chicago, IL, May 2001.Google Scholar
  42. 42.
    Marshall, A. G.; Wang, T.-C. L.; Ricca, T. L. Tailored Excitation for Fourier Transform Ion Cyclotron Resonance Mass Spectrometry. J. Am. Chem. Soc. 1985, 107, 7893–7897.CrossRefGoogle Scholar
  43. 43.
    Guan, S.; Marshall, A. G. Stored Waveform Inverse Fourier Transform (SWIFT) Ion Excitation in Trapped-Ion Mass Spectrometry: Theory and Applications. Int. J. Mass Spectrom. Ion Proc 1996, 157/158, 5–37.CrossRefGoogle Scholar
  44. 44.
    Wilcox, B. E.; Hendrickson, C. L.; Marshall, A. G. Improved Ion Extraction from a Linear Octopole Ion Trap: SIMION Analyis and Experimental Demonstration. J. Am. Soc. Mass Spectrom. 2002, 13, 1304–1312.CrossRefGoogle Scholar
  45. 45.
    Beu, S. C.; Laude, D. A. Jr Open Trapped Ion Cell Geometries for FT/ICR/MS. Int. J. Mass Spectrom. Ion Processes 1992, 112, 215–230.45.CrossRefGoogle Scholar
  46. 46.
    Håkansson, K.; Chalmers, M. J.; Quinn, J. P.; McFarland, M. A.; Hendrickson, C. L.; Marshall, A. G. Combined Electron Capture and Infrared Multiphoton Dissociation for Multistage MS/MS in a Fourier Transform Ion Cyclotron Resonance Mass Spectrometer. Anal. Chem. 2003, 75, 3256–3262.CrossRefGoogle Scholar
  47. 47.
    Ledford, E. B. Jr; Rempel, D. L.; Gross, M. L. Space Charge Effects in Fourier Transform Mass Spectrometry Mass Calibration. Anal. Chem. 1984, 56, 2744–2748.CrossRefGoogle Scholar
  48. 48.
    Shi, S. D.-H.; Drader, J. J.; Freitas, M. A.; Hendrickson, C. L.; Marshall, A. G. Comparison and Interconversion of the Two Most Common Frequency-to-Mass Calibration Functions for Fourier Transform Ion Cyclotron Resonance Mass Spectrometry. Int. J. Mass Spectrom 2000, 195/196, 591–598.CrossRefGoogle Scholar
  49. 49.
    Mansson, J. E.; Vanier, M. T.; Svennerholm, L. Changes in the Fatty Acid and Sphingosine Composition of the Major Gangliosides of Human Brain with Age. J. Neurochem. 1978, 30, 273–275.CrossRefGoogle Scholar
  50. 50.
    McLuckey, S. A.; Goeringer, D. E. Slow Heating Methods in Tandem Mass Spectrometry. J. Mass Spectrom. 1997, 35, 461–474.CrossRefGoogle Scholar
  51. 51.
    Kelleher, N. L.; Zubarev, R. A.; Bush, K.; Furie, B.; Furie, B. C.; McLafferty, F. W.; Walsh, C. T. Localization of Labile Posttranslational Modifications by Electron Capture Dissociation: The Case of γ-carboxyglutamic Acid. Anal. Chem. 1999, 71, 4250–4253.CrossRefGoogle Scholar
  52. 52.
    Leymarie, N.; Costello, C. E.; O’Connor, P. B. Electron Capture Dissociation Initiates a Free Radical Reaction Cascade. J. Am. Chem. Soc. 2003, 125, 8949–8958.CrossRefGoogle Scholar
  53. 53.
    Cooper, H. J.; Hudgins, R. R.; Håkansson, K.; Marshall, A. G. Secondary Fragmentation of Linear Peptides in Electron Capture Dissociation. Int. J. Mass Spectrom. 2003, 228, 723–728.CrossRefGoogle Scholar
  54. 54.
    Hughey, C. A.; Hendrickson, C. L.; Rodgers, R. P.; Marshall, A. G.; Qian, K. Kendrick Mass Defect Spectrum: A Compact Visual Analysis for Ultrahigh-Resolution Broadband Mass Spectra. Anal. Chem. 2001, 73, 4676–4681.CrossRefGoogle Scholar
  55. 55.
    Pomerantz, S.; McCloskey, J. Fractional Mass Values of Large Molecules. J. Mass Spectrom. 1987, 22, 251–253.CrossRefGoogle Scholar
  56. 56.
    Jones, J. J.; Stump, M. J.; Fleming, R. C.; Lay, J. O. Jr; Wilkins, C. L. Strategies and Data Analysis Techniques for Lipid and Phospholipid Chemistry Elucidation by Intact Cell MALDI-FTMS. J. Am. Soc. Mass Spectrom. 2004, 15, 1665–1674CrossRefGoogle Scholar

Copyright information

© American Society for Mass Spectrometry 2005

Authors and Affiliations

  • Melinda A. McFarland
    • 1
    • 4
  • Alan G. Marshall
    • 1
    • 4
  • Christopher L. Hendrickson
    • 1
    • 4
  • Carol L. Nilsson
    • 1
    • 3
  • Pam Fredman
    • 2
  • Jan-Eric Månsson
    • 2
  1. 1.Ion Cyclotron Resonance Program, National High Magnetic Field LaboratoryFlorida State UniversityTallahasseeUSA
  2. 2.Institute of Clinical NeuroscienceSahlgrenska University Hospital, Göteborg UniversityMölndalSweden
  3. 3.the Institute of Medical BiochemistryGöteborg UniversityGöteborgSweden
  4. 4.the Department of Chemistry and BiochemistryFlorida State UniversityTallahasseeUSA

Personalised recommendations