Abstract
A strategy for improving the sequencing of peptides by infrared multiphoton dissociation (IRMPD) in a linear ion trap mass spectrometer is described. We have developed an N-terminal derivatization reagent, 4-methylphosphonophenylisothiocyanate (PPITC), which allows the attachment of an IR-chromogenic phosphonite group to the N-terminus of peptides, thus enhancing their IRMPD efficiencies. After the facile derivatization process, the PPITC-modified peptides require shorter irradiation times for efficient IRMPD and yield extensive series of y ions, including those of low m/z that are not detected upon traditional CID. The resulting IRMPD mass spectra afford more complete sequence coverage for both model peptides and tryptic peptides from cytochrome c. We compare the effectiveness of this derivatization/IRMPD approach to that of a common N-terminal sulfonation reaction that utilizes 4-sulfophenylisothiocyanate (SPITC) in conjunction with CID and IRMPD.
Article PDF
Similar content being viewed by others
Avoid common mistakes on your manuscript.
References
Winston, R. L.; Fitzgerald, M. C. Mass Spectrometry as a Readout of Protein Structure and Function. Mass Spectrom. Rev. 1997, 16, 165–179.
Godovac-Zimmermann, J.; Brown, L. R. Perspectives for Mass Spectrometry and Functional Proteomics. Mass Spectrom. Rev. 2001, 20, 1–57.
Faux, M. C.; Scott, J. D. More on Target with Protein Phosphorylation: Conferring Specificity by Location. Trends Biochem. Sci. 1996, 21, 312–315.
Lill, J. Proteomic Tools for Quantitation by Mass Spectrometry. Mass Spectrom. Rev. 2003, 22, 182–194.
Regnier, F. E.; Riggs, L.; Zhang, R.; Xiong, L.; Liu, P.; Chakraborty, A.; Seeley, E.; Sioma, C.; Thompson, R. A. Comparative Proteomics Based on Stable Isotope Labeling and Affinity Selection. J. Mass Spectrom. 2002, 37, 133–145.
Sun, H.; Tonks, N. K. The Coordinated Action of Protein Tyrosine Phosphatases and Kinases in Cell Signaling. Trends Biochem. Sci. 1994, 19, 480–485.
Wysocki, V. H.; Resing, K. A.; Zhang, Q.; Cheng, G. Mass Spectrometry of Peptides and Proteins. Methods. 2005, 35, 211–222.
Koy, C.; Mikkat, S.; Raptakis, E.; Sutton, C.; Resch, M.; Tanaka, K.; Glocker, M. O. Matrix-Assisted Laser Desorption/Ionization-Quadrupole Ion Trap Time of Flight Mass Spectrometry Sequencing Resolves Structures of Unidentified Peptides Obtained by In-Gel Tryptic Digestion of Haptoglobin Derivatives from Human Plasma Proteomes. Proteomics. 2003, 3, 851–858.
Moyer, S. C.; Cotter, R. J.; Woods, A. S. Fragmentation of Phosphopeptides by Atmospheric Pressure MALDI and ESI/Ion Trap Mass Spectrometry. J. Am. Soc. Mass Spectrom. 2002, 13, 274–283.
Zubarev Roman, A. Electron-Capture Dissociation Tandem Mass Spectrometry. Curr. Opin. Biotechnol. 2004, 15, 12–16.
Syka, J. E. P.; Coon, J. J.; Schroeder, M. J.; Shabanowitz, J.; Hunt, D. F. Peptide and Protein Sequence Analysis by Electron Transfer Dissociation Mass Spectrometry. Proc. Natl. Acad. Sci. U.S.A. 2004, 101, 9528–9533.
Schwartz, J. C. High-Q Pulsed Fragmentation in Ion Traps. 2005, 2004941 653 6949743; 20040914.
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.
Wilson, J. J.; Brodbelt, J. S. Ultraviolet Photodissociation at 355 nm of Fluorescently Labeled Oligosaccharides. Anal. Chem. 2008, 80, 5186–5196.
Wilson, J. J.; Brodbelt, J. S. MS/MS Simplification by 355 nm Ultraviolet Photodissociation of Chromophore-Derivatized Peptides in a Quadrupole Ion Trap. Anal. Chem. 2007, 79, 7883–7892.
Wilson, J. J.; Brodbelt, J. S. Infrared Multiphoton Dissociation for Enhanced De Novo Sequence Interpretation of N-Terminal Sulfonated Peptides in a Quadrupole Ion Trap. Anal. Chem. 2006, 78, 6855–6862.
Payne, A. H.; Glish, G. L. Thermally Assisted Infrared Multiphoton Photodissociation in a Quadrupole Ion Trap. Anal. Chem. 2001, 73, 3542–3548.
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.
Crowe, M. C.; Brodbelt, J. S. Infrared Multiphoton Dissociation (IRMPD) and Collisionally Activated Dissociation of Peptides in a Quadrupole Ion Trap with Selective IRMPD of Phosphopeptides. J. Am. Soc. Mass Spectrom. 2004, 15, 1581–1592.
Eng, J. K.; McCormack, A. L.; Yates, J. R. III An Approach to Correlate Tandem Mass Spectral Data of Peptides with Amino Acid Sequences in a Protein Database. J. Am. Soc. Mass Spectrom. 1994, 5, 976–989.
Perkins, D. N.; Pappin, D. J. C.; Creasy, D. M.; Cottrell, J. S. Probability-Based Protein Identification by Searching Sequence Databases Using Mass Spectrometry Data. Electrophoresis. 1999, 20, 3551–3567.
Colorado, A.; Shen, J. X.; Brodbelt, J. Use of Infrared Multiphoton Photodissociation with SWIFT for Electrospray Ionization and Laser Desorption Applications in a Quadrupole Ion Trap Mass Spectrometer. Anal. Chem. 1996, 68, 4033–4043.
Crowe, M. C.; Brodbelt, J. S. Differentiation of Phosphorylated and Unphosphorylated Peptides by High-Performance Liquid Chromatography-Electrospray Ionization-Infrared Multiphoton Dissociation in a Quadrupole Ion Trap. Anal. Chem. 2005, 77, 5726–5734.
Crowe, M. C.; Brodbelt, J. S.; Goolsby, B. J.; Hergenrother, P. Characterization of Erythromycin Analogs by Collisionally Activated Dissociation and Infrared Multiphoton Dissociation in a Quadrupole Ion Trap. J. Am. Soc. Mass Spectrom. 2002, 13, 630–649.
Goolsby, B. J.; Brodbelt, J. S. Tandem Infrared Multiphoton Dissociation and Collisionally Activated Dissociation Techniques in a Quadrupole Ion Trap. Anal. Chem. 2001, 73, 1270–1276.
Hashimoto, Y.; Hasegawa, H.; Yoshinari, K.; Waki, I. Collisionally-Activated Infrared Multiphoton Dissociation in a Quadrupole Ion Trap Mass Spectrometer. Anal. Chem. 2003, 75, 420–425.
Keller, K. M.; Brodbelt, J. S. Collisionally Activated Dissociation and Infrared Multiphoton Dissociation of Oligonucleotides in a Quadrupole Ion Trap. Anal. Biochem. 2004, 326, 200–210.
Shen, J.; Brodbelt, J. S. Characterization of Ionophore-Metal Complexes by Infrared Multiphoton Photodissociation and Collisionally Activated Dissociation in a Quadrupole Ion Trap Mass Spectrometer. Analyst. 2000, 125, 641–650.
Wilson, J. J.; Kirkovits, G. J.; Sessler, J. L.; Brodbelt, J. S. Photodissociation of Noncovalent Peptide-Crown Ether Complexes. J. Am. Soc. Mass Spectrom. 2008, 19, 257–260.
Devakumar, A.; Mechref, Y.; Kang, P.; Novotny, M. V.; Reilly, J. P. Identification of Isomeric N-Glycan Structures by Mass Spectrometry with 157 nm Laser-Induced Photofragmentation. J. Am. Soc. Mass Spectrom. 2008, 19, 1027–1040.
Thompson, M. S.; Cui, W.; Reilly, J. P. Mass Spectrometry: Fragmentation of Singly Charged Peptide Ions by Photodissociation at l=157 nm. Angew. Chem. Int. Ed. 2004, 43, 4791–4794.
Ly, T.; Julian, R. R. Residue-Specific Radical-Directed Dissociation of Whole Proteins in the Gas Phase. J. Am. Chem. Soc. 2008, 130, 351–358.
Yoon, S. H.; Chung, Y. J.; Kim, M. S. Time-Resolved Photodissociation of Singly Protonated Peptides with an Arginine at the N-Terminus: A Statistical Interpretation. J. Am. Soc. Mass Spectrom. 2008, 19, 645–655.
Moon, J. H.; Shin, Y. S.; Cha, H. J.; Kim, M. S. Photodissociation at 193 nm of Some Singly Protonated Peptides and Proteins with m/z 2000-9000 Using a Tandem Time-of-Flight Mass Spectrometer Equipped with a Second Source for Delayed Extraction/Post-Acceleration of Product Ions. Rapid Commun. Mass Spectrom. 2007, 21, 359–368.
Yoon, S. H.; Kim, M. S. Development of a Time-Resolved Method for Photodissociation Mechanistic Study of Protonated Peptides: Use of a Voltage-Floated Cell in a Tandem Time-of-Flight Mass Spectrometer. J. Am. Soc. Mass Spectrom. 2007, 18, 1729–1739.
Krijgsveld, J.; Heck, A. J. R. Quantitative Proteomics by Metabolic Labeling with Stable Isotopes. Drug Discovery Today: Targets. 2004, 3, S11-S15.
Qu, J.; Straubinger, R. M. Improved Sensitivity for Quantification of Proteins Using Triply Charged Cleavable Isotope-Coded Affinity Tag Peptides. Rapid Commun. Mass Spectrom. 2005, 19, 2857–2864.
Yomota, C.; Ohnishi, Y. Determination of Biotin Following Derivatization with 2-Nitrophenylhydrazine by High-Performance Liquid Chromatography with On-Line UV Detection and Electrospray-Ionization Mass Spectrometry. J. Chromatogr. A. 2007, 1142, 231–235.
Li, H.; Schopfer, L.; Spaulding, R.; Thompson, C. M.; Lockridge, O. Identification of Organophosphate-Reactive Proteins by Tandem Mass Spectrometry. Chem. Biol. Interact. 2005, 157/158, 383–384.
Pikulski, M.; Hargrove, A.; Shabbir, S. H.; Anslyn, E. V.; Brodbelt, J. S. Sequencing and Characterization of Oligosaccharides Using Infrared Multiphoton Dissociation and Boronic Acid Derivatization in a Quadrupole Ion Trap. J. Am. Soc. Mass Spectrom. 2007, 18, 2094–2106.
Pikulski, M.; Wilson, J. J.; Aguilar, A.; Brodbelt, J. S. Amplification of Infrared Multiphoton Dissociation Efficiency in a Quadruple Ion Trap Using IR-Active Ligands. Anal. Chem. 2006, 78, 8512–8517.
Keough, T.; Youngquist, R. S.; Lacy, M. P. A Method for High-Sensitivity Peptide Sequencing Using Post-Source Decay Matrix-Assisted Laser Desorption Ionization. Proc. Natl. Acad. Sci. U.S.A. 1999, 96, 7131–7136.
Lee, Y. H.; Shin, J.-W.; Ryu, S.; Lee, S.-W.; Lee, C. H.; Lee, K. Enrichment of N-Terminal Sulfonated Peptides by a Water-Soluble Fullerene Derivative and its Applications to Highly Efficient Proteomics. Anal. Chim. Acta. 2006, 556, 140–144.
Leon, I. R.; Neves-Ferreira, A. G. C.; Valente, R. H.; Mota, E. M.; Lenzi, H. L.; Perales, J. Improved Protein Identification Efficiency by Mass Spectrometry Using N-Terminal Chemical Derivatization of Peptides from Angiostrongylus costaricensis, a Nematode with Unknown Genome. J. Mass Spectrom. 2007, 42, 1363–1374.
Wang, D.; Kalb Suzanne, R.; Cotter Robert, J. Improved Procedures for N-Terminal Sulfonation of Peptides for Matrix-Assisted Laser Desorption/Ionization Post-Source Decay Peptide Sequencing. Rapid Commun. Mass Spectrom. 2004, 18, 96–102.
Shin, J.-W.; Lee, Y. H.; Hwang, S.; Lee, S.-W. Observation of an Unusually Facile Fragmentation Pathway of Gas-Phase Peptide Ions: A Study on the Gas-Phase Fragmentation Mechanism and Energetics of Tryptic Peptides Modified with 4-Sulfophenyl Isothiocyanate (SPITC) and 4-Chlorosulfophenyl Isocyanate (SPC) and Their 18-Crown-6 Complexes. J. Mass Spectrom. 2007, 42, 380–388.
Oh, J. Y.; Moon, J. H.; Lee, Y. H.; Hyung, S.-W.; Lee, S.-W.; Kim, M. S. Photodissociation Tandem Mass Spectrometry at 266 nm of an Aliphatic Peptide Derivatized with Phenyl Isothiocyanate and 4-Sulfophenyl Isothiocyanate. Rapid Commun. Mass Spectrom. 2005, 19, 1283–1288.
Gardner, M. W.; Vasicek, L. A.; Shabbir, S.; Anslyn, E. V.; Brodbelt, J. S. Chromogenic Cross-Linker for the Characterization of Protein Structure by Infrared Multiphoton Dissociation Mass Spectrometry. Anal. Chem. 2008, 80, 4807–4819.
Flora, J. W.; Muddiman, D. C. Selective, Sensitive, and Rapid Phosphopeptide Identification in Enzymatic Digests Using ESI-FTICR-MS with Infrared Multiphoton Dissociation. Anal. Chem. 2001, 73, 3305–3311.
Flora, J. W.; Muddiman, D. C. Gas-Phase Ion Unimolecular Dissociation for Rapid Phosphopeptide Mapping by IRMPD in a Penning Ion Trap: An Energetically Favored Process. J. Am. Chem. Soc. 2002, 124, 6546–6547.
Flora, J. W.; Muddiman, D. C. Determination of the Relative Energies of Activation for the Dissociation of Aromatic vs: Aliphatic Phosphopeptides by ESI-FTICR-MS and IRMPD. J. Am. Soc. Mass Spectrom. 2004, 15, 121–127.
Jones, J. L.; Dongre, A. R.; Somogyi, A.; Wysocki, V. H. Sequence Dependence of Peptide Fragmentation Efficiency Curves Determined by Electrospray Ionization/Surface-Induced Dissociation Mass Spectrometry. J. Am. Chem. Soc. 1994, 116, 8368–8369.
Guillaume, E.; Panchaurd, A.; Affolter, M.; Desvergnes, V.; Kussmann, M. Differentially Isotope-Coded N-Terminal Protein Sulphonation: Combining Protein Identification and Quantification. Proteomics. 2006, 6, 2338–2349.
Lee, Y. H.; Kim, M.-S.; Choie, W.-S.; Min, H.-K.; Lee, S.-W. Highly Informative Proteome Analysis by Combining Improved N-Terminal Sulfonation for De Novo Peptide Sequencing and On-Line Capillary Reverse-Phase Liquid Chromatography/Tandem Mass Spectrometry. Proteomics. 2004, 4, 1684–1694.
Bunk, D. M.; Macfarlane, R. D. Derivatization to Enhance Sequence-Specific Fragmentation of Peptides and Proteins. Int. J. Mass Spectrom. Ion Processes. 1993, 126, 123–136.
Beardsley, R. L.; Karty, J. A.; Reilly, J. P. Enhancing the Intensities of Lysine-Terminated Tryptic Peptide Ions in Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry. Rapid Commun. Mass Spectrom. 2000, 14, 2147–2153.
Author information
Authors and Affiliations
Corresponding author
Additional information
Published online October 30, 2008
Electronic supplementary material
Rights and permissions
About this article
Cite this article
Vasicek, L.A., Wilson, J.J. & Brodbelt, J.S. Improved infrared multiphoton dissociation of peptides through N-terminal phosphonite derivatization. J Am Soc Mass Spectrom 20, 377–384 (2009). https://doi.org/10.1016/j.jasms.2008.10.016
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1016/j.jasms.2008.10.016