Methyl group migration during the fragmentation of singly charged ions of trimethyllysine-containing peptides: Precaution of using MS/MS of singly charged ions for interrogating peptide methylation



Core histones are susceptible to a range of post-translational modifications (PTMs), including acetylation, phosphorylation, methylation, and ubiquitination, which play important roles in the epigenetic control of gene expression. Here, we observed an unusual discrepancy between MALDI-MS/MS and ESI-MS/MS on the methylation of trimethyllysine-containing peptides with residues 9–17 from human histone H3 and residues 73–83 from yeast histone H3. It turned out that the discrepancy could be attributed to an unusual methyl group migration from the side chain of trimethyllysine to the C-terminal arginine residue during peptide fragmentation, and this methyl group transfer only occurred for singly charged ions, but not for doubly charged ions. The methyl group transfer argument received its support from the results on the studies of the fragmentation of the ESI- or MALDI-produced singly charged ions of several synthetic trimethyllysine-bearing peptides. The results presented in this study highlighted that caution should be exerted while MS/MS of singly charged ions is employed to interrogate the PTMs of trimethyllysine-containing peptides.

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  1. 1.
    Luger, K.; Mader, A. W.; Richmond, R. K.; Sargent, D. F.; Richmond, T. J. Crystal Structure of the Nucleosome Core Particle at 2.8 A Resolution. Nature 1997, 389, 251–260.CrossRefGoogle Scholar
  2. 2.
    Strahl, B. D.; Allis, C. D. The Language of Covalent Histone Modifications. Nature 2000, 403, 41–45.CrossRefGoogle Scholar
  3. 3.
    Jenuwein, T.; Allis, C. D. Translating the Histone Code. Science 2001, 293, 1074–1080.CrossRefGoogle Scholar
  4. 4.
    Klose, R. J.; Zhang, Y. Regulation of Histone Methylation by Demethylimination and Demethylation. Nat. Rev. Mol. Cell. Biol. 2007, 8, 307–318.CrossRefGoogle Scholar
  5. 5.
    Cocklin, R. R.; Wang, M. Identification of Methylation and Acetylation Sites on Mouse Histone H3 Using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight and Nanoelectrospray Ionization Tandem Mass Spectrometry. J. Protein Chem. 2003, 22, 327–334.CrossRefGoogle Scholar
  6. 6.
    Zhang, K.; Williams, K. E.; Huang, L.; Yau, P.; Siino, J. S.; Bradbury, E. M.; Jones, P. R.; Minch, M. J.; Burlingame, A. L. Histone acetylation and deacetylation: Identification of acetylation and methylation sites of HeLa histone H4 by mass spectrometry. Mol. Cell. Proteom. 2002, 1, 500–508.CrossRefGoogle Scholar
  7. 7.
    Gu, C.; Tsaprailis, G.; Breci, L.; Wysocki, V. H. Selective Gas-Phase Cleavage at the Peptide Bond C-Terminal to Aspartic Acid in Fixed-Charge Derivatives of Asp-Containing Peptides. Anal. Chem. 2000, 72, 5804–5813.CrossRefGoogle Scholar
  8. 8.
    Tsaprailis, G.; Nair, H.; Somogyi, A.; Wysocki, V. H.; Zhong, W. Q.; Futrell, J. H.; Summerfield, S. G.; Gaskell, S. J. Influence of Secondary Structure on the Fragmentation of Protonated Peptides. J. Am. Chem. Soc. 1999, 121, 5142–5154.CrossRefGoogle Scholar
  9. 9.
    Nair, H.; Wysocki, V. H. Are Peptides Without Basic Residues Protonated Primarily at the Amino Terminus? Int. J. Mass Spectrom. 1998, 174, 95–100.CrossRefGoogle Scholar
  10. 10.
    Dongre, A. R.; Somogyi, A.; Wysocki, V. H. Surface-Induced Dissociation: An Effective Tool to Probe Structure, Energetics, and Fragmentation Mechanisms of Protonated Peptides. J. Mass Spectrom. 1996, 31, 339–350.CrossRefGoogle Scholar
  11. 11.
    McCormack, A. L.; Somogyi, A.; Dongre, A. R.; Wysocki, V. H. Fragmentation of Protonated Peptides: Surface-Induced Dissociation in Conjunction with a Quantum Mechanical Approach. Anal. Chem. 1993, 65, 2859–2872.CrossRefGoogle Scholar
  12. 12.
    Tsaprailis, G.; Somogyi, A.; Nikolaev, E. N.; Wysocki, V. H. Refining the Model for Selective Cleavage at Acidic Residues in Arginine-Containing Protonated Peptides. Int. J. Mass Spectrom. 2000, 196, 467–479.CrossRefGoogle Scholar
  13. 13.
    de Maaijer-Gielbert, J.; Gu, C.; Somogyi, A.; Wysocki, V. H.; Kistemaker, P. G.; Weeding, T. L. Surface-Induced Dissociation of Singly and Multiply Protonated Polypropylenamine Dendrimers. J. Am. Soc. Mass Spectrom. 1999, 10, 414–422.CrossRefGoogle Scholar
  14. 14.
    Paizs, B.; Suhai, S. Fragmentation Pathways of Protonated Peptides. Mass Spectrom. Rev. 2005, 24, 508–548.CrossRefGoogle Scholar
  15. 15.
    Zhang, K.; Tang, H.; Huang, L.; Blankenship, J. W.; Jones, P. R.; Xiang, F.; Yau, P. M.; Burlingame, A. L. Identification of Acetylation and Methylation Sites of Histone H3 from Chicken Erythrocytes by High-Accuracy Matrix-Assisted Laser Desorption Ionization-Time-of-Flight, Matrix-Assisted Laser Desorption Ionization-Postsource Decay, and Nanoelectrospray Ionization Tandem Mass Spectrometry. Anal. Biochem. 2002, 306, 259–269.CrossRefGoogle Scholar
  16. 16.
    Edmondson, D. G.; Smith, M. M.; Roth, S. Y. Repression Domain of the Yeast Global Repressor Tup1 Interacts Directly with Histones H3 and H4. Genes Dev. 1996, 10, 1247–1259.CrossRefGoogle Scholar
  17. 17.
    Braunstein, M.; Rose, A. B.; Holmes, S. G.; Allis, C. D.; Broach, J. R. Transcriptional Silencing in Yeast is Associated with Reduced Nucleosome Acetylation. Genes Dev. 1993, 7, 592–604.CrossRefGoogle Scholar
  18. 18.
    Margueron, R.; Trojer, P.; Reinberg, D. The Key to Development: Interpreting the Histone Code? Curr. Opin. Genet. Dev. 2005, 15, 163–176.CrossRefGoogle Scholar
  19. 19.
    Stock, A.; Clarke, S.; Clarke, C.; Stock, J. N-Terminal Methylation of Proteins: Structure, Function, and Specificity. FEBS Lett. 1987, 220, 8–14.CrossRefGoogle Scholar
  20. 20.
    Miao, F.; Natarajan, R. Mapping Global Histone Methylation Patterns in the Coding Regions of Human Genes. Mol. Cell. Biol. 2005, 25, 4650–4661.CrossRefGoogle Scholar
  21. 21.
    Zhang, K.; Sridhar, V. V.; Zhu, J.; Kapoor, A.; Zhu, J. K. Distinctive Core Histone Post-Translational Modification Patterns in Arabidopsis thaliana. PLoS ONE 2007, 2, e1210.Google Scholar
  22. 22.
    Thorne, G. C.; Gaskell, S. J. Elucidation of Some Fragmentations of Small Peptides Using Sequential Mass Spectrometry on a Hybrid Instrument. Rapid Commun. Mass Spectrom. 1989, 3, 217–221.CrossRefGoogle Scholar
  23. 23.
    Zou, Y.; Wang, Y. Tandem Mass Spectrometry for the Examination of the Post-Translational Modifications of High-Mobility Group A1 Proteins: Symmetric and Asymmetric Dimethylation of Arg25 in HMGA1a Protein. Biochemistry 2005, 44, 6293–6301.CrossRefGoogle Scholar
  24. 24.
    Zou, Y.; Webb, K.; Perna, A. D.; Zhang, Q.; Clarke, S.; Wang, Y. A Mass Spectrometric Study on the In Vitro Methylation of HMGA1a and HMGA1b Proteins by PRMTs: Methylation Specificity, the Effect of Binding to AT-Rich Duplex DNA, and the Effect of C-Terminal Phosphorylation. Biochemistry 2007, 46, 7896–7906.CrossRefGoogle Scholar

Copyright information

© American Society for Mass Spectrometry 2009

Authors and Affiliations

  • Lei Xiong
    • 1
  • Liyan Ping
    • 1
  • Bifeng Yuan
    • 1
  • Yinsheng Wang
    • 1
  1. 1.Department of Chemistry-027University of California at RiverdaleRiverdaleUSA

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