Mass spectrometry approaches to study mammalian kinase and phosphatase associated proteins

Methods. 2012 Aug;57(4):400-8. doi: 10.1016/j.ymeth.2012.06.002. Epub 2012 Jun 16.


Reversible phosphorylation events regulate critical aspects of cellular biology by affecting protein conformation, cellular localization, enzymatic activity and associations with interaction partners. Kinases and phosphatases interact not only with their substrates but also with regulatory subunits and other proteins, including scaffolds. In recent years, affinity purification coupled to mass spectrometry (AP-MS) has proven to be a powerful tool to identify protein-protein interactions (PPIs) involving kinases and phosphatases. In this review we outline general considerations for successful AP-MS, and describe strategies that we have used to characterize the interactions of kinases and phosphatases in human cells.

Publication types

  • Research Support, Non-U.S. Gov't
  • Review

MeSH terms

  • Animals
  • Chromatography, Affinity
  • Humans
  • Phosphoprotein Phosphatases / chemistry
  • Phosphoprotein Phosphatases / isolation & purification
  • Phosphoprotein Phosphatases / metabolism*
  • Protein Binding
  • Protein Interaction Mapping / methods*
  • Protein Kinases / chemistry
  • Protein Kinases / isolation & purification
  • Protein Kinases / metabolism*
  • Proteolysis
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / isolation & purification
  • Recombinant Fusion Proteins / metabolism
  • Tandem Mass Spectrometry
  • Trypsin / chemistry


  • Recombinant Fusion Proteins
  • Protein Kinases
  • Phosphoprotein Phosphatases
  • Trypsin