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Measurement of Phosphorylated Peptides with Absolute Quantification

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Quantitative Proteomics by Mass Spectrometry

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1410))


Mass spectrometry, when coupled to on-line separation such as liquid chromatography or capillary electrophoresis, enables the identification and quantification of protein expression and post-translational modification changes under diverse conditions. To date most of the methods for mass spectrometry-based quantification have either provided relative quantification information (e.g., comparison to a selected condition) or utilized one-point calibration curves, or calibration curves in a different biological matrix. Although these quantitative methods have been used to generate insight into the differences between biological samples, additional biological insight could be gained by accurately measuring the absolute quantity of selected proteins and protein modifications. To address this challenge, we have developed the MARQUIS (Multiplex Absolute Regressed Quantification with Internal Standards) method, designed to provide absolute quantification for potentially hundreds of peptides across multiple samples in a single analysis, using a multi-point internal calibration curve derived from synthetic, isotopically distinct standard peptides.

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  1. Morris M, Chi A, Melas I et al (2014) Phosphoproteomics in drug discovery. Drug Discov Today 19:425–432

    Article  CAS  PubMed  Google Scholar 

  2. Jones R, Gordus A, Krall J et al (2005) A quantitative protein interaction network for the ErbB receptor using protein microarrays. Nature 439:168–174

    Article  PubMed  Google Scholar 

  3. Liang S, Xu Z, Xu X et al (2012) Quantitative proteomics for cancer biomarker discovery. Comb Chem High Throughput Screen 15:221–231

    Article  CAS  PubMed  Google Scholar 

  4. Wasinger V, Zeng M, Yau Y (2013) Current status and advances in quantitative proteomic mass spectrometry. Int J Proteomics 2013:1–12

    Article  Google Scholar 

  5. Wolf-Yadlin A, Kumar N, Zhang Y et al (2006) Effects of HER2 overexpression on cell signaling networks governing proliferation and migration. Mol Syst Biol 2:1–15

    Article  Google Scholar 

  6. Gerber S, Rush J, Stemman O et al (2003) Absolute quantification of proteins and phosphoproteins from cell lysates by tandem MS. Proc Natl Acad Sci U S A 100:6940–6945

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  7. Curran T, Zhang Y, Ma D et al (2015) MARQUIS: a multiplex method for absolute quantification of peptides and posttranslational modifications. Nat Commun 6:1–11

    Article  Google Scholar 

  8. Yarden Y (2001) The EGFR family and its ligands in human cancer: signalling mechanisms and therapeutic opportunities. Eur J Cancer 37:S3–S8

    Article  CAS  PubMed  Google Scholar 

  9. Gan H, Cvrljevic A, Johns T (2013) The epidermal growth factor receptor variant III (EGFRvIII): where wild things are altered. FEBS J 280:5350–5370

    Article  CAS  PubMed  Google Scholar 

  10. Schulze W, Deng L, Mann M (2005) Phosphotyrosine interactome of the ErbB-receptor kinase family. Mol Syst Biol 1:42–54

    Article  Google Scholar 

  11. Moser K, White F (2006) Phosphoproteomic analysis of rat liver by high capacity IMAC and LC-MS/MS. J Proteome Res 5:98–104

    Article  CAS  PubMed  Google Scholar 

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Correspondence to Forest M. White .

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© 2016 Springer Science+Business Media New York

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Reddy, R.J., Curran, T.G., Zhang, Y., White, F.M. (2016). Measurement of Phosphorylated Peptides with Absolute Quantification. In: Sechi, S. (eds) Quantitative Proteomics by Mass Spectrometry. Methods in Molecular Biology, vol 1410. Humana Press, New York, NY.

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3522-2

  • Online ISBN: 978-1-4939-3524-6

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