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Mass spectrometry–based proteomics turns quantitative

Abstract

The field of proteomics is built on technologies to analyze large numbers of proteins—ideally the entire proteome—in the same experiment. Mass spectrometry (MS) has been successfully used to characterize proteins in complex mixtures, but results so far have largely been qualitative. Two recently developed methodologies offer the opportunity to obtain quantitative proteomic information. Comparing the signals from the same peptide under different conditions yields a rough estimate of relative protein abundance between two proteomes. Alternatively, and more accurately, peptides are labeled with stable isotopes, introducing a predictable mass difference between peptides from two experimental conditions. Stable isotope labels can be incorporated 'post-harvest', by chemical approaches or in live cells through metabolic incorporation. This isotopic handle facilitates direct quantification from the mass spectra. Using these quantitative approaches, precise functional information as well as temporal changes in the proteome can be captured by MS.

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Figure 1: Extracting quantitative data from mass spectra.
Figure 2: PCP to determine the centrosomal proteome.
Figure 3: Stages of incorporation of stable isotope labels and their impact on quantitative accuracy.
Figure 4: Quantification of proteins with extensive sequence identity.
Figure 5: Identification of specific bait-prey interactions in affinity-precipitation experiments.
Figure 6: Temporal changes in the nucleolar proteome upon transcriptional inhibition.

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References

  1. Wilkins, M.R. et al. Progress with proteome projects: why all proteins expressed by a genome should be identified and how to do it. Biotechnol. Genet. Eng. Rev. 13, 19–50 (1996).

    CAS  PubMed  Google Scholar 

  2. Anderson, N.L. & Anderson, N.G. Proteome and proteomics: new technologies, new concepts, and new words. Electrophoresis 19, 1853–1861 (1998).

    CAS  PubMed  Google Scholar 

  3. Corthals, G.L., Wasinger, V.C., Hochstrasser, D.F. & Sanchez, J.C. The dynamic range of protein expression: a challenge for proteomic research. Electrophoresis 21, 1104–1115 (2000).

    CAS  PubMed  Google Scholar 

  4. Aebersold, R. & Mann, M. Mass spectrometry–based proteomics. Nature 422, 198–207 (2003).

    CAS  PubMed  Google Scholar 

  5. Peng, J. & Gygi, S.P. Proteomics: the move to mixtures. J. Mass Spectrom. 36, 1083–1091 (2001).

    CAS  PubMed  Google Scholar 

  6. Brunet, S. et al. Organelle proteomics: looking at less to see more. Trends Cell Biol. 13, 629–638 (2003).

    CAS  PubMed  Google Scholar 

  7. Garcia, B.A., Shabanowitz, J. & Hunt, D.F. Analysis of protein phosphorylation by mass spectrometry. Methods 35, 256–264 (2005).

    CAS  PubMed  Google Scholar 

  8. Kuster, B., Schirle, M., Mallick, P. & Aebersold, R. Scoring proteomes with proteotypic peptide probes. Nat. Rev. Mol. Cell Biol. 6, 577–583 (2005).

    CAS  PubMed  Google Scholar 

  9. Lill, J. Proteomic tools for quantitation by mass spectrometry. Mass Spectrom. Rev. 22, 182–194 (2003).

    CAS  PubMed  Google Scholar 

  10. Sadygov, R.G., Cociorva, D. & Yates, J.R. Large-scale database searching using tandem mass spectra: Looking up the answer in the back of the book. Nat Methods 1, 195–202 (2004).

    CAS  PubMed  Google Scholar 

  11. Wysocki, V.H., Resing, K.A., Zhang, Q. & Cheng, G. Mass spectrometry of peptides and proteins. Methods 35, 211–222 (2005).

    CAS  PubMed  Google Scholar 

  12. Julka, S. & Regnier, F. Quantification in proteomics through stable isotope coding: a review. J. Proteome Res. 3, 350–363 (2004).

    CAS  PubMed  Google Scholar 

  13. Leitner, A. & Lindner, W. Current chemical tagging strategies for proteome analysis by mass spectrometry. J. Chromatogr. B Analyt. Technol. Biomed. Life Sci. 813, 1–26 (2004).

    CAS  PubMed  Google Scholar 

  14. Olsen, J.V., Ong, S.E. & Mann, M. Trypsin cleaves exclusively C-terminal to arginine and lysine residues. Mol. Cell. Proteomics 3, 608–614 (2004).

    CAS  PubMed  Google Scholar 

  15. Allet, N. et al. In vitro and in silico processes to identify differentially expressed proteins. Proteomics 4, 2333–2351 (2004).

    CAS  PubMed  Google Scholar 

  16. Rappsilber, J., Ishihama, I., Foster, L.J., Mittler, G. & Mann, M. in Proceedings of the 51st American Society for Mass Spectrometry Conference on Mass Spectrometry (American Society for Mass Spectrometry, Montreal 2003).

    Google Scholar 

  17. Steen, H. & Mann, M. The ABC's (and XYZ's) of peptide sequencing. Nat. Rev. Mol. Cell Biol. 5, 699–711 (2004).

    CAS  PubMed  Google Scholar 

  18. Rappsilber, J., Ryder, U., Lamond, A.I. & Mann, M. Large-scale proteomic analysis of the human spliceosome. Genome Res. 12, 1231–1245 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  19. Sanders, S.L., Jennings, J., Canutescu, A., Link, A.J. & Weil, P.A. Proteomics of the eukaryotic transcription machinery: identification of proteins associated with components of yeast TFIID by multidimensional mass spectrometry. Mol. Cell. Biol. 22, 4723–4738 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  20. Ishihama, Y. et al. Exponentially modified protein abundance index (emPAI) for estimation of absolute protein amount in proteomics by the number of sequenced peptides per protein. Mol. Cell Proteomics, advance online publication 2005 06 14 (10.174/mcp.M500061-MCP200).

  21. Andersen, J.S. et al. Proteomic characterization of the human centrosome by protein correlation profiling. Nature 426, 570–574 (2003).

    CAS  PubMed  Google Scholar 

  22. Chelius, D. & Bondarenko, P.V. Quantitative profiling of proteins in complex mixtures using liquid chromatography and mass spectrometry. J. Proteome Res. 1, 317–323 (2002).

    CAS  PubMed  Google Scholar 

  23. Gillette, M.A., Mani, D.R. & Carr, S.A. Place of pattern in proteomic biomarker discovery. J. Proteome Res. 4, 1143–1154 (2005).

    CAS  PubMed  Google Scholar 

  24. Carr, S.A., Huddleston, M.J. & Annan, R.S. Selective detection and sequencing of phosphopeptides at the femtomole level by mass spectrometry. Anal. Biochem. 239, 180–192 (1996).

    CAS  PubMed  Google Scholar 

  25. Verma, R. et al. Phosphorylation of Sic1p by G1 Cdk required for its degradation and entry into S phase. Science 278, 455–460 (1997).

    CAS  PubMed  Google Scholar 

  26. Steen, H., Jebanathirajah, J.A., Springer, M. & Kirschner, M.W. Stable isotope-free relative and absolute quantitation of protein phosphorylation stoichiometry by MS. Proc. Natl. Acad. Sci. USA 102, 3948–3953 (2005).

    CAS  PubMed  Google Scholar 

  27. De Leenheer, A.P. & Thienpont, L.M. Applications of isotope dilution-mass spectrometry in clinical chemistry, pharmcokinetics, and toxicology. Mass Spectrom. Rev. 11, 249–307 (1992).

    CAS  Google Scholar 

  28. Wulfkuhle, J.D., Liotta, L.A. & Petricoin, E.F. Proteomic applications for the early detection of cancer. Nat. Rev. Cancer 3, 267–275 (2003).

    CAS  PubMed  Google Scholar 

  29. Zhang, R. & Regnier, F.E. Minimizing resolution of isotopically coded peptides in comparative proteomics. J. Proteome Res. 1, 139–147 (2002).

    CAS  PubMed  Google Scholar 

  30. Kusmierz, J.J., Sumrada, R. & Desiderio, D.M. Fast atom bombardment mass spectrometric quantitative analysis of methionine-enkephalin in human pituitary tissues. Anal. Chem. 62, 2395–2400 (1990).

    CAS  PubMed  Google Scholar 

  31. Stemmann, O., Zou, H., Gerber, S.A., Gygi, S.P. & Kirschner, M.W. Dual inhibition of sister chromatid separation at metaphase. Cell 107, 715–726 (2001).

    CAS  PubMed  Google Scholar 

  32. Kuhn, E. et al. Quantification of C-reactive protein in the serum of patients with rheumatoid arthritis using multiple reaction monitoring mass spectrometry and 13C-labeled peptide standards. Proteomics 4, 1175–1186 (2004).

    CAS  PubMed  Google Scholar 

  33. Hopfgartner, G. et al. Triple quadrupole linear ion trap mass spectrometer for the analysis of small molecules and macromolecules. J. Mass Spectrom. 39, 845–855 (2004).

    CAS  PubMed  Google Scholar 

  34. Kirkpatrick, D.S., Gerber, S.A. & Gygi, S.P. The absolute quantification strategy: a general procedure for the quantification of proteins and post-translational modifications. Methods 35, 265–273 (2005).

    CAS  PubMed  Google Scholar 

  35. Havlis, J. & Shevchenko, A. Absolute quantification of proteins in solutions and in polyacrylamide gels by mass spectrometry. Anal. Chem. 76, 3029–3036 (2004).

    CAS  PubMed  Google Scholar 

  36. Aebersold, R. Constellations in a cellular universe. Nature 422, 115–116 (2003).

    CAS  PubMed  Google Scholar 

  37. Beynon, R.J., Doherty, M.K., Pratt, J.M. & Gaskell, S.J. Multiplexed absolute quantification in proteomics using artificial QCAT proteins of concatenated signatures. Nature Methods 2, 587–589 (2005).

    CAS  PubMed  Google Scholar 

  38. Rose, K. et al. A new mass-spectrometric C-terminal sequencing technique finds a similarity between γ-interferon and α 2-interferon and identifies a proteolytically clipped γ-interferon that retains full antiviral activity. Biochem. J. 215, 273–277 (1983).

    CAS  PubMed  PubMed Central  Google Scholar 

  39. Mirgorodskaya, O.A. et al. Quantitation of peptides and proteins by matrix-assisted laser desorption/ionization mass spectrometry using (18)O-labeled internal standards. Rapid Commun. Mass Spectrom. 14, 1226–1232 (2000).

    CAS  PubMed  Google Scholar 

  40. Yao, X., Freas, A., Ramirez, J., Demirev, P.A. & Fenselau, C. Proteolytic 18O labeling for comparative proteomics: model studies with two serotypes of adenovirus. Anal. Chem. 73, 2836–2842 (2001).

    CAS  PubMed  Google Scholar 

  41. Rao, K.C., Palamalai, V., Dunlevy, J.R. & Miyagi, M. Lys-N catalyzed 18O labeling for comparative proteomics: Application to cytokines/LPS treated human retinal pigment epithelium cell line. Mol. Cell Proteomics, advance online publication 2005 07 05 (10.1074/mcp.M500150-MCP200).

  42. Yao, X., Afonso, C. & Fenselau, C. Dissection of proteolytic 18O labeling: endoprotease-catalyzed 16O-to-18O exchange of truncated peptide substrates. J. Proteome Res. 2, 147–152 (2003).

    CAS  PubMed  Google Scholar 

  43. Bantscheff, M., Dumpelfeld, B. & Kuster, B. Femtomol sensitivity post-digest (18)O labeling for relative quantification of differential protein complex composition. Rapid Commun. Mass Spectrom. 18, 869–876 (2004).

    CAS  PubMed  Google Scholar 

  44. Staes, A. et al. Global differential non-gel proteomics by quantitative and stable labeling of tryptic peptides with oxygen-18. J. Proteome Res. 3, 786–791 (2004).

    CAS  PubMed  Google Scholar 

  45. Gygi, S.P. et al. Quantitative analysis of complex protein mixtures using isotope-coded affinity tags. Nat. Biotechnol. 17, 994–999 (1999).

    CAS  PubMed  Google Scholar 

  46. Hansen, K.C. et al. Mass spectrometric analysis of protein mixtures at low levels using cleavable 13C-isotope-coded affinity tag and multidimensional chromatography. Mol. Cell. Proteomics 2, 299–314 (2003).

    CAS  PubMed  Google Scholar 

  47. Li, J., Steen, H. & Gygi, S.P. Protein profiling with cleavable isotope-coded affinity tag (cICAT) reagents: the yeast salinity stress response. Mol. Cell. Proteomics 2, 1198–1204 (2003).

    CAS  PubMed  Google Scholar 

  48. Oda, Y. et al. Quantitative chemical proteomics for identifying candidate drug targets. Anal. Chem. 75, 2159–2165 (2003).

    CAS  PubMed  Google Scholar 

  49. Olsen, J.V. et al. HysTag–a novel proteomic quantification tool applied to differential display analysis of membrane proteins from distinct areas of mouse brain. Mol. Cell. Proteomics 3, 82–92 (2004).

    CAS  PubMed  Google Scholar 

  50. Nielsen, P.A. et al. Proteomic mapping of brain plasma membrane proteins. Mol. Cell. Proteomics 4, 402–408 (2005).

    CAS  PubMed  Google Scholar 

  51. Goodlett, D.R. et al. Differential stable isotope labeling of peptides for quantitation and de novo sequence derivation. Rapid Commun. Mass Spectrom. 15, 1214–1221 (2001).

    CAS  PubMed  Google Scholar 

  52. Ficarro, S.B. et al. Phosphoproteome analysis by mass spectrometry and its application to Saccharomyces cerevisiae. Nat. Biotechnol. 20, 301–305 (2002).

    CAS  PubMed  Google Scholar 

  53. Salomon, A.R. et al. Profiling of tyrosine phosphorylation pathways in human cells using mass spectrometry. Proc. Natl. Acad. Sci. USA 100, 443–448 (2003).

    CAS  PubMed  Google Scholar 

  54. Brill, L.M. et al. Robust phosphoproteomic profiling of tyrosine phosphorylation sites from human T cells using immobilized metal affinity chromatography and tandem mass spectrometry. Anal. Chem. 76, 2763–2772 (2004).

    CAS  PubMed  Google Scholar 

  55. Munchbach, M., Quadroni, M., Miotto, G. & James, P. Quantitation and facilitated de novo sequencing of proteins by isotopic N-terminal labeling of peptides with a fragmentation-directing moiety. Anal. Chem. 72, 4047–4057 (2000).

    CAS  PubMed  Google Scholar 

  56. Ji, J. et al. Strategy for qualitative and quantitative analysis in proteomics based on signature peptides. J. Chromatogr. B Biomed. Sci. Appl. 745, 197–210 (2000).

    CAS  PubMed  Google Scholar 

  57. Schmidt, A., Kellermann, J. & Lottspeich, F. A novel strategy for quantitative proteomics using isotope-coded protein labels. Proteomics 5, 4–15 (2005).

    CAS  PubMed  Google Scholar 

  58. Zhang, X., Jin, Q.K., Carr, S.A. & Annan, R.S. N-Terminal peptide labeling strategy for incorporation of isotopic tags: a method for the determination of site-specific absolute phosphorylation stoichiometry. Rapid Commun. Mass Spectrom. 16, 2325–2332 (2002).

    CAS  PubMed  Google Scholar 

  59. Ross, P.L. et al. Multiplexed protein quantitation in Saccharomyces cerevisiae using amine-reactive isobaric tagging reagents. Mol. Cell. Proteomics 3, 1154–1169 (2004).

    CAS  PubMed  Google Scholar 

  60. Langen, H., Fountoulakis, M., Evers, S., Wipf, B. & Berndt, P. in From Genome to Proteome, 3rd Siena 2D Electrophoresis Meeting (Wiley-VCH, Weinheim, Germany, Siena, 1998).

    Google Scholar 

  61. Oda, Y., Huang, K., Cross, F.R., Cowburn, D. & Chait, B.T. Accurate quantitation of protein expression and site-specific phosphorylation. Proc. Natl. Acad. Sci. USA 96, 6591–6596 (1999).

    CAS  PubMed  Google Scholar 

  62. Conrads, T.P. et al. Quantitative analysis of bacterial and mammalian proteomes using a combination of cysteine affinity tags and 15N-metabolic labeling. Anal. Chem. 73, 2132–2139 (2001).

    CAS  PubMed  Google Scholar 

  63. Krijgsveld, J. et al. Metabolic labeling of C. elegans and D. melanogaster for quantitative proteomics. Nat. Biotechnol. 21, 927–931 (2003).

    CAS  PubMed  Google Scholar 

  64. Wu, C.C., MacCoss, M.J., Howell, K.E., Matthews, D.E. & Yates, J.R.I. Metabolic labeling of mammalian organisms with stable isotopes for quantitative proteomic analysis. Anal. Chem. 76, 4951–4959 (2004).

    CAS  PubMed  Google Scholar 

  65. Ong, S.E. et al. Stable isotope labeling by amino acids in cell culture, SILAC, as a simple and accurate approach to expression proteomics. Mol. Cell. Proteomics 1, 376–386 (2002).

    CAS  PubMed  Google Scholar 

  66. Chen, X., Smith, L.M. & Bradbury, E.M. Site-specific mass tagging with stable isotopes in proteins for accurate and efficient protein identification. Anal. Chem. 72, 1134–1143 (2000).

    CAS  PubMed  Google Scholar 

  67. Veenstra, T.D., Martinovic, S., Anderson, G.A., Pasa-Tolic, L. & Smith, R.D. Proteome analysis using selective incorporation of isotopically labeled amino acids. J. Am. Soc. Mass Spectrom. 11, 78–82 (2000).

    CAS  PubMed  Google Scholar 

  68. Zhu, H., Pan, S., Gu, S., Bradbury, E.M. & Chen, X. Amino acid residue specific stable isotope labeling for quantitative proteomics. Rapid Commun. Mass Spectrom. 16, 2115–2123 (2002).

    CAS  PubMed  Google Scholar 

  69. Jiang, H. & English, A.M. Quantitative analysis of the yeast proteome by incorporation of isotopically labeled leucine. J. Proteome Res. 1, 345–350 (2002).

    CAS  PubMed  Google Scholar 

  70. Ong, S.E., Kratchmarova, I. & Mann, M. Properties of 13C-substituted arginine in stable isotope labeling by amino acids in cell culture (SILAC). J. Proteome Res. 2, 173–181 (2003).

    CAS  PubMed  Google Scholar 

  71. Foster, L.J., De Hoog, C.L. & Mann, M. Unbiased quantitative proteomics of lipid rafts reveals high specificity for signaling factors. Proc. Natl. Acad. Sci. USA 100, 5813–5818 (2003).

    CAS  PubMed  Google Scholar 

  72. Ibarrola, N., Kalume, D.E., Gronborg, M., Iwahori, A. & Pandey, A. A proteomic approach for quantitation of phosphorylation using stable isotope labeling in cell culture. Anal. Chem. 75, 6043–6049 (2003).

    CAS  PubMed  Google Scholar 

  73. Andersen, J.S. et al. Nucleolar proteome dynamics. Nature 433, 77–83 (2005).

    CAS  PubMed  Google Scholar 

  74. Ibarrola, N., Molina, H., Iwahori, A. & Pandey, A. A novel proteomic approach for specific identification of tyrosine kinase substrates using [13C]tyrosine. J. Biol. Chem. 279, 15805–15813 (2004).

    CAS  PubMed  Google Scholar 

  75. Ong, S.E., Mittler, G. & Mann, M. Identifying and quantifying in vivo methylation sites by heavy methyl SILAC. Nat. Methods 1, 119–126 (2004).

    CAS  PubMed  Google Scholar 

  76. Scott, L., Lamb, J., Smith, S. & Wheatley, D.N. Single amino acid (arginine) deprivation: rapid and selective death of cultured transformed and malignant cells. Br. J. Cancer 83, 800–810 (2000).

    CAS  PubMed  PubMed Central  Google Scholar 

  77. Gruhler, A. et al. Quantitative phosphoproteomics applied to the yeast pheromone signaling pathway. Mol. Cell. Proteomics 4, 310–327 (2005).

    CAS  PubMed  Google Scholar 

  78. Gehrmann, M.L., Hathout, Y. & Fenselau, C. Evaluation of metabolic labeling for comparative proteomics in breast cancer cells. J. Proteome Res. 3, 1063–1068 (2004).

    CAS  PubMed  Google Scholar 

  79. Amanchy, R., Kalume, D.E. & Pandey, A. Stable isotope labeling with amino acids in cell culture (SILAC) for studying dynamics of protein abundance and posttranslational modifications. Sci. STKE 267, l2 (2005).

    Google Scholar 

  80. Ong, S.E. et al. Stable isotope labeling by amino acids in cell culture (SILAC) for quantitative proteomics. in Handbook of Cell Biology 3rd edn. Vol. IV (ed. Celis, J.) (Academic Press, San Diego, 2005).

    Google Scholar 

  81. Ishihama, Y. et al. Quantitative mouse brain proteomics using culture-derived isotope tags as internal standards. Nat. Biotechnol. 23, 617–621 (2005).

    CAS  PubMed  Google Scholar 

  82. Schweitzer, B. & Kingsmore, S.F. Measuring proteins on microarrays. Curr. Opin. Biotechnol. 13, 14–19 (2002).

    CAS  PubMed  Google Scholar 

  83. Zhu, H., Bilgin, M. & Snyder, M. Proteomics. Annu. Rev. Biochem. 72, 783–812 (2003).

    CAS  PubMed  Google Scholar 

  84. Rappsilber, J. & Mann, M. What does it mean to identify a protein in proteomics? Trends Biochem. Sci. 27, 74–78 (2002).

    CAS  PubMed  Google Scholar 

  85. Nesvizhskii, A.I. & Aebersold, R. Interpretation of shotgun proteomics data: The protein inference problem. Mol. Cell Proteomics, advance online publication 2005 07 11 (10.1074/mcp.R500012-MCP200).

  86. Han, D.K., Eng, J., Zhou, H. & Aebersold, R. Quantitative profiling of differentiation-induced microsomal proteins using isotope-coded affinity tags and mass spectrometry. Nat. Biotechnol. 19, 946–951 (2001).

    CAS  PubMed  PubMed Central  Google Scholar 

  87. MacCoss, M.J., Wu, C.C., Liu, H., Sadygov, R. & Yates, J.R., III. A correlation algorithm for the automated quantitative analysis of shotgun proteomics data. Anal. Chem. 75, 6912–6921 (2003).

    CAS  PubMed  Google Scholar 

  88. Schulze, W.X. & Mann, M. A novel proteomic screen for peptide-protein interactions. J. Biol. Chem. 279, 10756–10764 (2004).

    CAS  PubMed  Google Scholar 

  89. Shiio, Y. et al. Quantitative proteomic analysis of Myc oncoprotein function. EMBO J. 21, 5088–5096 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  90. Everley, P.A., Krijgsveld, J., Zetter, B.R. & Gygi, S.P. Quantitative cancer proteomics: stable isotope labeling with amino acids in cell culture (SILAC) as a tool for prostate cancer research. Mol. Cell. Proteomics 3, 729–735 (2004).

    CAS  PubMed  Google Scholar 

  91. Gu, S. et al. Large-scale quantitative proteomic study of PUMA-induced apoptosis using two-dimensional liquid chromatography-mass spectrometry coupled with amino acid-coded mass tagging. J. Proteome Res. 3, 1191–1200 (2004).

    CAS  PubMed  Google Scholar 

  92. Ho, Y. et al. Systematic identification of protein complexes in Saccharomyces cerevisiae by mass spectrometry. Nature 415, 180–183 (2002).

    CAS  PubMed  Google Scholar 

  93. Gavin, A.C. et al. Functional organization of the yeast proteome by systematic analysis of protein complexes. Nature 415, 141–147 (2002).

    CAS  PubMed  Google Scholar 

  94. Blagoev, B. et al. A proteomics strategy to elucidate functional protein-protein interactions applied to EGF signaling. Nat. Biotechnol. 21, 315–318 (2003).

    CAS  PubMed  Google Scholar 

  95. Ranish, J.A. et al. The study of macromolecular complexes by quantitative proteomics. Nat. Genet. 33, 349–355 (2003).

    CAS  PubMed  Google Scholar 

  96. Blagoev, B., Ong, S.E., Kratchmarova, I. & Mann, M. Temporal analysis of phosphotyrosine-dependent signaling networks by quantitative proteomics. Nat. Biotechnol. 22, 1139–1145 (2004).

    CAS  PubMed  Google Scholar 

  97. Zhang, Y. et al. Time-resolved mass spectrometry of tyrosine phosphorylatiopn sites in the EGF receptor signaling network reveals dynamic modules. Mol. Cell Proteomics, advance online publication 2005 06 11 (10.1074/mcp.M500089-MCP200).

  98. Pratt, J.M. et al. Dynamics of protein turnover, a missing dimension in proteomics. Mol. Cell. Proteomics 1, 579–591 (2002).

    CAS  PubMed  Google Scholar 

  99. Saghatelian, A. & Cravatt, B.F. Assignment of protein function in the postgenomic era. Nat Chem. Biol. 1, 130–142 (2005).

    CAS  PubMed  Google Scholar 

  100. Verhelst, S.H. & Bogyo, M. Chemical proteomics applied to target identification and drug discovery. Biotechniques 38, 175–177 (2005).

    CAS  PubMed  Google Scholar 

  101. Sechi, S. & Chait, B.T. Modification of cysteine residues by alkylation. A tool in peptide mapping and protein identification. Anal. Chem. 70, 5150–5158 (1998).

    CAS  PubMed  Google Scholar 

  102. Shen, M. et al. Isolation and isotope labeling of cysteine- and methionine-containing tryptic peptides: application to the study of cell surface proteolysis. Mol. Cell. Proteomics 2, 315–324 (2003).

    CAS  PubMed  Google Scholar 

  103. Sebastiano, R., Citterio, A., Lapadula, M. & Righetti, P.G. A new deuterated alkylating agent for quantitative proteomics. Rapid Commun. Mass Spectrom. 17, 2380–2386 (2003).

    CAS  PubMed  Google Scholar 

  104. Pasquarello, C., Sanchez, J.C., Hochstrasser, D.F. & Corthals, G.L. N-t-butyliodoacetamide and iodoacetanilide: two new cysteine alkylating reagents for relative quantitation of proteins. Rapid Commun. Mass Spectrom. 18, 117–127 (2004).

    CAS  PubMed  Google Scholar 

  105. Zhou, H., Ranish, J.A., Watts, J.D. & Aebersold, R. Quantitative proteome analysis by solid-phase isotope tagging and mass spectrometry. Nat. Biotechnol. 20, 512–515 (2002).

    CAS  PubMed  Google Scholar 

  106. Qiu, Y., Sousa, E.A., Hewick, R.M. & Wang, J.H. Acid-labile isotope-coded extractants: a class of reagents for quantitative mass spectrometric analysis of complex protein mixtures. Anal. Chem. 74, 4969–4979 (2002).

    CAS  PubMed  Google Scholar 

  107. Shi, Y. et al. A simple solid phase mass tagging approach for quantitative proteomics. J. Proteome Res. 3, 104–111 (2004).

    CAS  PubMed  Google Scholar 

  108. Shi, Y., Xiang, R., Horvath, C. & Wilkins, J.A. Quantitative analysis of membrane proteins from breast cancer cell lines BT474 and MCF7 using multistep solid phase mass tagging and 2D LC/MS. J. Proteome Res. 4, 1427–1433 (2005).

    CAS  PubMed  Google Scholar 

  109. Thompson, A. et al. Tandem mass tags: a novel quantification strategy for comparative analysis of complex protein mixtures by MS/MS. Anal. Chem. 75, 1895–1904 (2003).

    CAS  PubMed  Google Scholar 

  110. Wang, S. & Regnier, F.E. Proteomics based on selecting and quantifying cysteine containing peptides by covalent chromatography. J. Chromatogr. A. 924, 345–357 (2001).

    CAS  PubMed  Google Scholar 

  111. Che, F.Y. & Fricker, L.D. Quantitation of neuropeptides in Cpe(fat)/Cpe(fat) mice using differential isotopic tags and mass spectrometry. Anal. Chem. 74, 3190–3198 (2002).

    CAS  PubMed  Google Scholar 

  112. Mason, D.E. & Liebler, D.C. Quantitative analysis of modified proteins by LC-MS/MS of peptides labeled with phenyl isocyanate. J. Proteome Res. 2, 265–272 (2003).

    CAS  PubMed  Google Scholar 

  113. Lee, Y.H., Han, H., Chang, S.B. & Lee, S.W. Isotope-coded N-terminal sulfonation of peptides allows quantitative proteomic analysis with increased de novo peptide sequencing capability. Rapid Commun. Mass Spectrom. 18, 3019–3027 (2004).

    CAS  PubMed  Google Scholar 

  114. Hoang, V.M. et al. Quantitative proteomics employing primary amine affinity tags. J. Biomol. Tech. 14, 216–223 (2003).

    PubMed  PubMed Central  Google Scholar 

  115. Hsu, J.L., Huang, S.Y., Chow, N.H. & Chen, S.H. Stable-isotope dimethyl labeling for quantitative proteomics. Anal. Chem. 75, 6843–6852 (2003).

    CAS  PubMed  Google Scholar 

  116. Cagney, G. & Emili, A. De novo peptide sequencing and quantitative profiling of complex protein mixtures using mass-coded abundance tagging. Nat. Biotechnol. 20, 163–170 (2002).

    CAS  PubMed  Google Scholar 

  117. Brancia, F.L., Openshaw, M.E. & Kumashiro, S. Investigation of the electrospray response of lysine-, arginine-, and homoarginine-terminal peptide mixtures by liquid chromatography/mass spectrometry. Rapid Commun. Mass Spectrom. 16, 2255–2259 (2002).

    CAS  PubMed  Google Scholar 

  118. Brancia, F.L., Montgomery, H., Tanaka, K. & Kumashiro, S. Guanidino labeling derivatization strategy for global characterization of peptide mixtures by liquid chromatography matrix-assisted laser desorption/ionization mass spectrometry. Anal. Chem. 76, 2748–2755 (2004).

    CAS  PubMed  Google Scholar 

  119. Beardsley, R.L. & Reilly, J.P. Quantitation using enhanced signal tags: a technique for comparative proteomics. J. Proteome Res. 2, 15–21 (2003).

    CAS  PubMed  Google Scholar 

  120. Peters, E.C., Horn, D.M., Tully, D.C. & Brock, A. A novel multifunctional labeling reagent for enhanced protein characterization with mass spectrometry. Rapid Commun. Mass Spectrom. 15, 2387–2392 (2001).

    CAS  PubMed  Google Scholar 

  121. Syka, J.E. et al. Novel linear quadrupole ion trap/FT mass spectrometer: performance characterization and use in the comparative analysis of histone H3 post-translational modifications. J. Proteome Res. 3, 621–626 (2004).

    CAS  PubMed  Google Scholar 

  122. Kuyama, H. et al. An approach to quantitative proteome analysis by labeling tryptophan residues. Rapid Commun. Mass Spectrom. 17, 1642–1650 (2003).

    CAS  PubMed  Google Scholar 

  123. Goshe, M.B. et al. Phosphoprotein isotope-coded affinity tag approach for isolating and quantitating phosphopeptides in proteome-wide analyses. Anal. Chem. 73, 2578–2586 (2001).

    CAS  PubMed  Google Scholar 

  124. Qian, W.J. et al. Phosphoprotein isotope-coded solid-phase tag approach for enrichment and quantitative analysis of phosphopeptides from complex mixtures. Anal. Chem. 75, 5441–5450 (2003).

    CAS  PubMed  Google Scholar 

  125. Wells, L. et al. Mapping sites of O-GlcNAc modification using affinity tags for serine and threonine post-translational modifications. Mol. Cell. Proteomics 1, 791–804 (2002).

    CAS  PubMed  Google Scholar 

  126. Amoresano, A., Marino, G., Cirulli, C. & Quemeneur, E. Mapping phosphorylation sites: a new strategy based on the use of isotopically labelled DTT and mass spectrometry. Eur. J. Mass Spectrom. (Chichester, Eng) 10, 401–412 (2004).

    CAS  Google Scholar 

  127. Vosseller, K. et al. Quantitative analysis of both protein expression and serine / threonine post-translational modifications through stable isotope labeling with dithiothreitol. Proteomics 5, 388–398 (2005).

    CAS  PubMed  Google Scholar 

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Acknowledgements

We gratefully acknowledge our colleagues at the Broad Institute of MIT and Harvard, the Center for Experimental BioInformatics at University of Southern Denmark and the Max Planck Institute for Biochemistry for useful discussions and support. We thank S.A. Carr, B. Küster, L.J. Foster and F. White for critical comments.

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Correspondence to Shao-En Ong or Matthias Mann.

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Ong, SE., Mann, M. Mass spectrometry–based proteomics turns quantitative. Nat Chem Biol 1, 252–262 (2005). https://doi.org/10.1038/nchembio736

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