Identification and Quantitation of Signal Molecule-Dependent Protein Phosphorylation

  • Arnoud Groen
  • Ludivine Thomas
  • Kathryn Lilley
  • Claudius Marondedze
Part of the Methods in Molecular Biology book series (MIMB, volume 1016)


Phosphoproteomics is a fast-growing field that aims at characterizing phosphorylated proteins in a cell or a tissue at a given time. Phosphorylation of proteins is an important regulatory mechanism in many cellular processes. Gel-free phosphoproteome technique involving enrichment of phosphopeptide coupled with mass spectrometry has proven to be invaluable to detect and characterize phosphorylated proteins. In this chapter, a gel-free quantitative approach involving 15N metabolic labelling in combination with phosphopeptide enrichment by titanium dioxide (TiO2) and their identification by MS is described. This workflow can be used to gain insights into the role of signalling molecules such as cyclic nucleotides on regulatory networks through the identification and quantification of responsive phospho(proteins).

Key words

Gel-free proteomics Mass spectrometry Phosphoproteomics Quantitation TiO2 enrichment 


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Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Arnoud Groen
    • 1
  • Ludivine Thomas
    • 2
  • Kathryn Lilley
    • 1
  • Claudius Marondedze
    • 1
  1. 1.Department of Biochemistry, Cambridge Centre for Proteomics, Cambridge Systems Biology CentreUniversity of CambridgeCambridgeUK
  2. 2.Division of Chemical and Life Sciences and EngineeringKing Abdullah University of Science and TechnologyThuwalSaudi Arabia

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