Journal of the American Society for Mass Spectrometry

, Volume 18, Issue 12, pp 2146–2161

Electron capture in charge-tagged peptides. Evidence for the role of excited electronic states

  • Julia Chamot-Rooke
  • Christian Malosse
  • Gilles Frison
  • František Tureček


Electron capture dissociation (ECD) was studied with doubly charged dipeptide ions that were tagged with fixed-charge tris-(2,4,6-trimethoxyphenyl)phosphonium-methylenecarboxamido (TMPP-ac) groups. Dipeptides GK, KG, AK, KA, and GR were each selectively tagged with one TMPP-ac group at the N-terminal amino group while the other charge was introduced by protonation at the lysine or arginine side-chain groups to give (TMPP-ac-peptide + H)2+ ions by electrospray ionization. Doubly tagged peptide derivatives were also prepared from GK, KG, AK, and KA in which the fixed-charge TMPP-ac groups were attached to the N-terminal and lysine side-chain amino groups to give (TMPP-ac-peptide-ac-TMPP)2+ dications by electrospray. ECD of (TMPP-ac-peptide + H)2+ resulted in 72% to 84% conversion to singly charged dissociation products while no intact charge-reduced (TMPP-ac-dipeptide + H) ions were detected. The dissociations involved loss of H, formation of (TMPP + H)+, and N-cα bond cleavages giving TMPP-CH2CONH2+ (c0) and c1 fragments. In contrast, ECD of (TMPP-ac-peptide-ac-TMPP)2+ resulted in 31% to 40% conversion to dissociation products due to loss of neutral TMPP molecules and 2,4,6-trimethoxyphenyl radicals. No peptide backbone cleavages were observed for the doubly tagged peptide ions. Ab initio and density functional theory calculations for (Ph3P-ac-GK + H)2+ and (H3P-ac-GK + H)2+ analogs indicated that the doubly charged ions contained the lysine side-chain NH3+ group internally solvated by the COOH group. The distance between the charge-carrying phosphonium and ammonium atoms was calculated to be 13.1–13.2 Å in the most stable dication conformers. The intrinsic recombination energies of the TMPP+-ac and (GK + H)+ moieties, 2. 7 and 3. 15 eV, respectively, indicated that upon electron capture the ground electronic states of the (TMPP-ac-peptide + H) ions retained the charge in the TMPP group. Ground electronic state (TMPP-ac-GK + H) ions were calculated to spontaneously isomerize by lysine H-atom transfer to the COOH group to form dihydroxycarbinyl radical intermediates with the retention of the charged TMPP group. These can trigger cleavages of the adjacent N-Cα bonds to give rise to the c1 fragment ions. However, the calculated transition-state energies for GK and GGK models suggested that the ground-state potential energy surface was not favorable for the formation of the abundant c0 fragment ions. This pointed to the involvement of excited electronic states according to the Utah-Washington mechanism of ECD.

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  1. 1.
    Tureček, F. Transient Intermediates of Chemical Reactions by Neutralization-Reionization Mass Spectrometry. Top. Curr. Chem. 2003, 225, 77–129.CrossRefGoogle Scholar
  2. 2.
    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
  3. 3.
    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
  4. 4.
    Rowe, B. R.; Mitchell, J. B.; Canosa, A. Dissociative Recombination. Theory, Experiment, and Applications; Plenum Press: New York, 1993; pp. 1–11.CrossRefGoogle Scholar
  5. 5.
    Cooper, H. J.; Hakansson, K.; Marshall, A. G. The role of electron capture dissociation in biomolecular analysis. Mass Spectrom. Rev. 2005, 24, 201–222.CrossRefGoogle Scholar
  6. 6.(a)
    Zubarev, R. A.; Horn, D. M.; Fridriksson, E. K.; Kelleher, N. L.; Kruger, N. A.; Lewis, M. A.; Carpenter, B. K.; McLafferty, F. W.. Anal. Chem. 2000, 72, 563–573.CrossRefGoogle Scholar
  7. 6.(b)
    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
  8. 6.(c)
    Fung, Y. M. E.; Chan, T.-W. D. Experimental and Theoretical Investigations of the Loss of Amino Acid Side Chains in Electron Capture Dissociation of Model Peptides. J. Am. Soc. Mass Spectrom. 2005, 16, 1523–1535.CrossRefGoogle Scholar
  9. 6.(d)
    O’Connor, P. B.; Lin, C.; Cournoyer, J. J.; Pittman, J. L.; Belyayev, M.; Budnik, B. A. Long-Lived: Electron Capture Dissociation Product Ions Experience Radical Migration via Hydrogen Abstraction. J. Am. Soc. Mass Spectrom. 2006, 17, 576–585.CrossRefGoogle Scholar
  10. 6.(e)
    Lin, C.; Cournoyer, J. J.; O’Connor, P. B. Use of a Double Resonance Electron Capture Dissociation Experiment to Probe Fragment Intermediate Lifetimes. J. Am. Soc. Mass Spectrom. 2006, 17, 1605–1615.CrossRefGoogle Scholar
  11. 7.(a)
    Konishi, H.; Yokotake, Y.; Ishibashi, T. Theoretical Study on the Electron Capture Dissociation Correlated with Proton Transfer Processes. J. Mass Spectrom. Soc. Jpn. 2002, 50, 222–225.CrossRefGoogle Scholar
  12. 7.(b)
    Bakken, V.; Helgaker, T.; Uggerud, E. Models of Fragmentations Induced by Electron Attachment to Protonated Peptides. Eur. J. Mass Spectrom. 2004, 10, 625–638.CrossRefGoogle Scholar
  13. 8.
    Tureček, F.; Syrstad, E. A. Mechanism and Energetics of Intramolecular Hydrogen Transfer Atom Transfer in Amide and Peptide Radicals and Cation-Radicals. J. Am. Chem. Soc. 2003, 125, 3353–3369.CrossRefGoogle Scholar
  14. 9.
    Tureček, F. N-Cα Bond Dissociation Energies and Kinetics in Amide and Peptide Radicals: Is the Dissociation a Non-Ergodic Process? J. Am. Chem. Soc. 2003, 125, 5954–5963.CrossRefGoogle Scholar
  15. 10.
    Tureček, F.; Syrstad, E. A.; Seymour, J. L.; Chen, X.; Yao, C. Peptide Cation-Radicals: A Computational Study of the Competition between Peptide N-Cα Bond Cleavage and Loss of the Side Chain in the [GlyPhe-NH2 + 2H] Cation Radical. J. Mass Spectrom. 2003, 38, 1093–1104.CrossRefGoogle Scholar
  16. 11.
    Chen, X.; Tureček, F. The Arginine Anomaly: Arginine Radicals are Poor Hydrogen Atom Donors in Electron Transfer Induced Dissociations. J. Am. Chem. Soc. 2006, 128, 12520–12530.CrossRefGoogle Scholar
  17. 12.
    Yao, C.; Fung, Y. M. E.; Tureček, F. Histidine Radicals. Proceedings of the UPPCON IV Conference, Hong Kong, December, 2006.Google Scholar
  18. 13.
    Yao, C.; Syrstad, E. A.; Tureček, F. Electron Transfer to Protonated β-Alanine N-Methylamide in the Gas Phase: An Experimental and Computational Study of Dissociation Energetics and Mechanisms. J. Phys. Chem. A. 2007, 111, 4167–4180.CrossRefGoogle Scholar
  19. 14.
    Syrstad, E. A.; Tureček, F. Toward a General Mechanism of Electron Capture Dissociation. J. Am. Soc. Mass Spectrom. 2005, 16, 208–224.CrossRefGoogle Scholar
  20. 15.(a)
    Sobczyk, M.; Anusiewicz, I.; Berdys-Kochanska, J.; Sawicka, A.; Skurski, P.; Simons, J. Coulomb-Assisted Dissociative Electron Attachment: Application to a Model Peptide. J. Phys. Chem. A. 2005, 109, 250–258.CrossRefGoogle Scholar
  21. 15.(b)
    Anusiewicz, I.; Berdys-Kochanska, J.; Skurski, P.; Simons, J. Simulating Electron Transfer Attachment to a Positively Charged Model Peptide. J. Phys. Chem. A. 2006, 110, 1261–1266.CrossRefGoogle Scholar
  22. 15.(c)
    Skurski, P.; Sobczyk, M.; Jakowski, J.; Simons, J. Possible Mechanisms for Protecting N-Cα bonds in helical peptides from electron capture (or transfer) dissociation. Int. J. Mass Spectrom. 2007, 265, 197–212.CrossRefGoogle Scholar
  23. 16.
    Huang, Z.-H.; Wu, J.; Roth, K. D. W.; Yang, Y.; Gage, D. A.; Watson, J. T. A Picomole-Scale Method for Charge Derivatization of Peptides for Sequence Analysis by Mass Spectrometry. Anal. Chem. 1997, 69, 137–144.CrossRefGoogle Scholar
  24. 17.
    Sadagopan, N.; Watson, J. T. Investigation of the Tris(Trimethoxyphenyl)Phosphonium Acetyl Charge Derivatives of Peptides by Electrospray Ionization Mass Spectrometry and Tandem Mass Spectrometry. J. Am. Soc. Mass Spectrom. 1999, 11, 107–119.CrossRefGoogle Scholar
  25. 18.
    Chamot-Rooke, J.; van der Rest, G.; Dalleu, A.; Bay, A.; Lemoine, J. The Combination of Electron Capture Dissociation and Fixed Charge Derivatization Increases Sequence Coverage for O-Glycosylated and O-Phosphorylated Peptides. J. Am. Soc. Mass Spectrom. 2007, 18, 1405–1413.CrossRefGoogle Scholar
  26. 19.
    Gunawardena, H. P.; Gorenstein, L.; Erickson, D. E.; Xia, Y.; McLuckey, S. A. Electron Transfer Dissociation of Multiply Protonated and Fixed Charge Disulfide Linked Polypeptides. Int. J. Mass Spectrom. 2007, 265, 130–138.CrossRefGoogle Scholar
  27. 20.
    Frisch, M. J.; Trucks, G. W.; Schlegel, H. B.; Scuseria, G. E.; Robb, M. A.; Cheeseman, J. R.; Montgomery, J. A. Jr.; Vreven, T.; Kudin, K. N.; Burant, J. C.; Millam, J. M.; Iyengar, S. S.; Tomasi, J.; Barone, V.; Mennucci, B.; Cossi, M.; Scalmani, G.; Rega, N.; Petersson, G. A.; Nakatsuji, H.; Hada, M.; Ehara, M.; Toyota, K.; Fukuda, R.; Hasegawa, J.; Ishida, M.; Nakajima, T.; Honda, Y.; Kitao, O.; Nakai, H.; Klene, M.; Li, X.; Knox, J. E.; Hratchian, H. P.; Cross, J. B.; Adamo, C.; Jaramillo, J.; Gomperts, R.; Stratmann, R. E.; Yazyev, O.; Austn, A. J.; Cammi, R.; Pomelli, C.; Ochterski, J. W.; Ayala, P. Y.; Morokuma, K.; Voth, G. A.; Salvador, P.; Dannenberg, J. J.; Zakrzewski, V. G.; Dapprich, S.; Daniels, A. D.; Strain, M. C.; Farkas, O.; Malick, D. K.; Rabuck, A. D.; Raghavachari, K.; Foresman, J. B.; Ortiz, J. V.; Cui, Q.; Baboul, A. G.; Clifford, S.; Cioslowski, J.; Stefanov, B. B.; Liu, G.; Liashenko, A.; Piskorz, P.; Komaromi, I.; Martin, R. L.; Fox, D. J.; Keith, T.; Al-Laham, M. A.; Peng, C. Y.; Nanayakkara, A.; Challacombe, M.; Gill, P. M. W.; Johnson, B.; Chen, W.; Wong, M. W.; Gonzalez, C.; Pople, J. A. Gaussian 03, Revision B-05; Gaussian, Inc.: Pittsburgh PA, 2003.Google Scholar
  28. 21.(a)
    Becke, A. D. A New Mixing of Hartree-Fock and Local Density-Functional Theories. J. Chem. Phys. 1993, 98, 1372–1377.CrossRefGoogle Scholar
  29. 21.(b)
    Becke, A. D. Density Functional Thermochemistry: III. The Role of Exact Exchange. J. Chem. Phys. 1993, 98, 5648–5652.CrossRefGoogle Scholar
  30. 22.
    Yao, C.; Tureček, F. Hypervalent Ammonium Radicals: Competitive N-C and N-H Bond Dissociations in Methylammonium and Ethylammonium. Phys. Chem., Chem. Phys. 2005, 7, 912–920.CrossRefGoogle Scholar
  31. 23.
    Rauhut, G.; Pulay, P. Transferable Scaling Factors for Density Functional Derived Vibrational Force Fields. J. Phys. Chem. 1995, 99, 3093–3100.CrossRefGoogle Scholar
  32. 24.
    Møller, C.; Plesset, M. S. A Note on an Approximation Treatment for Many-Electron Systems. Phys. Rev. 1934, 46, 618–622.CrossRefGoogle Scholar
  33. 25.(a)
    Schlegel, H. B. Potential Energy Curves Using Unrestricted Moller-Plesset Perturbation Theory with Spin Annihilation. J. Chem. Phys. 1986, 84, 4530–4534.CrossRefGoogle Scholar
  34. 25.(b)
    Mayer, I. Spin-Projected UHF Method: IV. Comparison of Potential Curves Given by Different One-Electron Methods. Adv. Quantum. Chem. 1980, 12, 189–262.CrossRefGoogle Scholar
  35. 26.(a)
    Tureček, F. Proton Affinity of Dimethyl Sulfoxide and Relative Stabilities of C2H6OS Molecules and C2H7OS+ Ions: A Comparative G2(MP2) ab Initio and Density Functional Theory Study. J. Phys. Chem. A. 1998, 102, 4703–4713.CrossRefGoogle Scholar
  36. 26.(b)
    Tureček, F.; Wolken, J. K. Dissociation Energies and Kinetics of Aminopyrimidinium Radicals by ab Initio and Density Functional Theory. J. Phys. Chem. A. 1999, 103, 1905–1912.Google Scholar
  37. 26.(c)
    Tureček, F.; Polašek, M.; Frank, A. J.; Sadilek, M. Transient Hydrogen Atom Adducts to Disulfides Formation and Energetics. J. Am. Chem. Soc. 2000, 122, 2361–2370.CrossRefGoogle Scholar
  38. 26.(d)
    Polašek, M.; Tureček, F. Hydrogen Atom Adducts to Nitrobenzene Formation of the Phenylnitronic Radical in the Gas Phase and Energetics of Wheland Intermediates. J. Am. Chem. Soc. 2000, 122, 9511–9524.CrossRefGoogle Scholar
  39. 26.(e)
    Rablen, P. R. Is the Acetate Anion Stabilized by Resonance or Electrostatics?: A Systematic Structural Comparison. J. Am. Chem. Soc. 2000, 122, 357–368.CrossRefGoogle Scholar
  40. 26.(f)
    Rablen, P. R. Computational Analysis of the Solvent Effect on the Barrier to Rotation about the Conjugated C-N Bond in Methyl N,N-Dimethylcarbamate. J. Org. Chem. 2000, 65, 7930–7937.CrossRefGoogle Scholar
  41. 26.(g)
    Rablen, P. R.; Bentrup, K. H. Are the Enolates of Amides and Esters Stabilized by Electrostatics? J. Am. Chem. Soc. 2003, 125, 2142–2147.CrossRefGoogle Scholar
  42. 26.(h)
    Hirama, M.; Tokosumi, T.; Ishida, T.; Aihara, J. Possible Molecular Hydrogen Formation Mediated by the Inner and Outer Carbon Atoms of Typical PAH cations. Chem. Phys. 2004, 305, 307–316.CrossRefGoogle Scholar
  43. 27.
    Čižek, J.; Paldus, J.; Šroubkova, L. Cluster Expansion Analysis for Delocalized Systems. Int. J. Quantum. Chem. 1969, 3, 149–167.CrossRefGoogle Scholar
  44. 28.
    Purvis, G. D. III; Bartlett, R. J. A Full Coupled-Cluster Singles and Doubles model: The Inclusion of Disconnected Triples. J. Chem. Phys. 1982, 76, 1910–1918.CrossRefGoogle Scholar
  45. 29.(a)
    Dunning, T. H. Jr. Gaussian Basis Sets for Use in Correlated Molecular Calculations: I. The Atoms Boron through Neon and Hydrogen. J. Chem. Phys. 1989, 90, 1007–1023.CrossRefGoogle Scholar
  46. 29.(b)
    Woon, D. E.; Dunning, T. H. Jr. Gaussian Basis Sets for Use in Correlated Molecular Calculations: III. The Atoms Aluminum through Argon. J. Chem. Phys. 1993, 98, 1358–1371.CrossRefGoogle Scholar
  47. 30.
    Stratmann, R. E.; Scuseria, G. E.; Frisch, M. J. An Efficient Implementation of Time-Dependent Density Functional Theory for the Calculation of Excitation Energies of Large Molecules. J. Chem. Phys. 1998, 109, 8218–8224.CrossRefGoogle Scholar
  48. 31.
    Reed, A. E.; Weinstock, R. B.; Weinhold, F. Natural Population Analysis. J. Chem. Phys. 1985, 83, 735–746.CrossRefGoogle Scholar
  49. 32.(a)
    Roepstorff, P.; Fohlman, J. Proposal for a Common Nomenclature for Sequence Ions in Mass Spectra of Peptides. Biomed. Mass Spectrom. 1984, 11, 601.CrossRefGoogle Scholar
  50. 32.(b)
    Johnson, R. S.; Martin, S. A.; Biemann, K. Collision-Induced Fragmentation of (M + H)+ ions of peptides: Side Chain Specific Sequence Ions. Int. J. Mass Spectrom. Ion Processes. 1988, 86, 137–154.CrossRefGoogle Scholar
  51. 33.
    McLafferty, F. W.; Tureček, F. Interpretation of Mass Spectra, 4th ed.; University Science Books: Mill Valley, CA, 1993; 19–26.Google Scholar
  52. 34.(a)
    Clareboudt, J.; Baten, W.; Geise, H.; Claeys, M. Structural Characterization of Mono- and Biphosphonium Salts by Fast Atom Bombardment Mass Spectrometry and Tandem Mass Spectrometry. Org. Mass Spectrom. 1993, 28, 71–82.CrossRefGoogle Scholar
  53. 34.(b)
    NIST Standard Reference Database Number 69, June 2005 Release, Scholar
  54. 35.(a)
    Gustafson, S. M.; Cramer, C. J. Ab Initio Conformational and Stereopermutational Analyses of Phosphoranyl Radicals HP(OR)3 and P(OR)4 [R = H or CH3]. J. Phys. Chem. 1995, 99, 2267–2277.CrossRefGoogle Scholar
  55. 35.(b)
    Tureček, F.; Gu, M.; Hop, C. E. C. A. Franck-Condon Dominated Chemistry Formation and Dissociations of Tetrahydroxyphosphoranyl Radicals Following Femtosecond Reduction of Their Cations in the Gas Phase. J. Phys. Chem. 1995, 99, 2278–2291.CrossRefGoogle Scholar
  56. 35.(c)
    Cramer, C. J. The Fluorophosphoranyl Series: Theoretical Insights into Relative Stabilities and Localization of Spin. J. Am. Chem. Soc. 1991, 113, 2439–2447.CrossRefGoogle Scholar
  57. 35.(d)
    Cramer, C. J. Computational Studies of Open-Shell Phosphorus Oxyacids: Pt. 3. Theoretical Rotation, Pseudorotation, and Pseudoinversion Barriers for the Hydroxyphosphoranyl Radical. J. Am. Chem. Soc. 1990, 112, 7965–7972.CrossRefGoogle Scholar
  58. 36.(a)
    Siegbahn, P. E. M.; Blomberg, M. R. A.; Crabtree, R. H. Hydrogen Transfer in the Presence of Amino Acid Radicals. Theor. Chem. Acc. 1997, 97, 289–300.CrossRefGoogle Scholar
  59. 36.(b)
    Siegbahn, P. E. M.; Eriksson, L.; Himo, F.; Pavlov, M. Hydrogen Atom Transfer in Ribonucleotide Reductase (RNR). J. Phys. Chem. B. 1998, 102, 10622–10629.CrossRefGoogle Scholar
  60. 36.(c)
    Cujier, R. L.; Nocera, D. G. Proton-Coupled Electron Transfer. Annu. Rev. Phys. Chem. 1998, 49, 337–369.CrossRefGoogle Scholar
  61. 37.
    Harrison, A. G. The Gas-Phase Basicities and Proton Affinities of Amino Acids and Peptides. Mass Spectrom. Rev. 1997, 16, 201–217.CrossRefGoogle Scholar

Copyright information

© American Society for Mass Spectrometry 2007

Authors and Affiliations

  • Julia Chamot-Rooke
    • 1
  • Christian Malosse
    • 1
  • Gilles Frison
    • 1
  • František Tureček
    • 2
  1. 1.Laboratoire des Mécanismes Réactionnels, Department of Chemistry, Ecole PolytechniqueCNRSPalaiseauFrance
  2. 2.Department of ChemistryUniversity of WashingtonSeattleUSA

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