Quantifying protein interaction dynamics by SWATH mass spectrometry: application to the 14-3-3 system

Nat Methods. 2013 Dec;10(12):1246-53. doi: 10.1038/nmeth.2703. Epub 2013 Oct 27.


Protein complexes and protein interaction networks are essential mediators of most biological functions. Complexes supporting transient functions such as signal transduction processes are frequently subject to dynamic remodeling. Currently, the majority of studies on the composition of protein complexes are carried out by affinity purification and mass spectrometry (AP-MS) and present a static view of the system. For a better understanding of inherently dynamic biological processes, methods to reliably quantify temporal changes of protein interaction networks are essential. Here we used affinity purification combined with sequential window acquisition of all theoretical spectra (AP-SWATH) mass spectrometry to study the dynamics of the 14-3-3β scaffold protein interactome after stimulation of the insulin-PI3K-AKT pathway. The consistent and reproducible quantification of 1,967 proteins across all stimulation time points provided insights into the 14-3-3β interactome and its dynamic changes following IGF1 stimulation. We therefore establish AP-SWATH as a tool to quantify dynamic changes in protein-complex interaction networks.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 14-3-3 Proteins / chemistry*
  • Chromatography, Affinity / methods
  • Computational Biology / methods
  • Gene Library
  • HEK293 Cells
  • Humans
  • Mass Spectrometry / methods*
  • Mechanistic Target of Rapamycin Complex 1
  • Mechanistic Target of Rapamycin Complex 2
  • Multiprotein Complexes / chemistry
  • Peptides / chemistry
  • Phosphatidylinositol 3-Kinases / chemistry
  • Protein Binding
  • Protein Interaction Mapping / methods*
  • Proteins / chemistry
  • Proteomics / methods
  • Signal Transduction
  • TOR Serine-Threonine Kinases / chemistry
  • Time Factors


  • 14-3-3 Proteins
  • Multiprotein Complexes
  • Peptides
  • Proteins
  • Mechanistic Target of Rapamycin Complex 1
  • Mechanistic Target of Rapamycin Complex 2
  • TOR Serine-Threonine Kinases