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Autophagy pp 341-357 | Cite as

Identification and Regulation of Multimeric Protein Complexes in Autophagy via SILAC-Based Mass Spectrometry Approaches

  • Stéphanie Kaeser-Pebernard
  • Britta Diedrich
  • Jörn DengjelEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1880)

Abstract

Mass spectrometry (MS)-based identification and characterization of protein complexes is becoming a prerequisite for in-depth biochemical analyses of intracellular processes. Here, we describe two state-of-the-art MS-based approaches to characterize protein-protein interactions and multi-protein complexes involved in autophagy in mammalian cells. The combination of affinity purification (AP)-MS, which identifies binary protein-protein interactions, with size-exclusion chromatography (SEC)-protein correlation profiling (PCP), which helps monitor protein complex assemblies, is a powerful tool to acquire a full overview of the interlinkage and regulation of novel multi-protein complexes that might play a role in autophagy.

Key words

Macroautophagy Autophagy Starvation SILAC AP-MS SEC-PCP-SILAC Proteomics Mass spectrometry 

Notes

Acknowledgments

This work was generously supported by the Swiss National Science Foundation (grants 31003A-166482/1, 316030-177088 ), by the Novartis Foundation for Medical-Biological Research (grant 16C221), by TRANSAUTOPHAGY (COST Action CA15138), and by the Alexander von Humboldt Foundation.

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Stéphanie Kaeser-Pebernard
    • 1
  • Britta Diedrich
    • 2
  • Jörn Dengjel
    • 1
    • 3
    Email author
  1. 1.Department of BiologyUniversity of FribourgFribourgSwitzerland
  2. 2.Agilent TechnologiesWaldbronnGermany
  3. 3.Department of DermatologyMedical Center—University of FreiburgFreiburgGermany

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