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Global Identification of Caspase Substrates Using PROTOMAP (Protein Topography and Migration Analysis Platform)

  • Melissa M. Dix
  • Gabriel M. Simon
  • Benjamin F. Cravatt
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1133)

Abstract

Delineation of the natural substrate scope of proteases is important for understanding the functions of proteolytic pathways in physiology and disease. Herein we describe the protocol for PROTOMAP, a technique that combines SDS-PAGE with tandem mass spectrometry to globally identify shifts in protein migration indicative of proteolytic processing. When applied to cells undergoing apoptosis, this unbiased global method provides a snapshot of the topography and magnitude of proteolytic events associated with programmed cell death.

Key words

SDS-PAGE In-gel digestion Mass spectrometry Spectral counting PROTOMAP Peptograph Substrate identification 

References

  1. 1.
    Dix MM, Simon GM, Cravatt BF (2008) Global mapping of the topography and magnitude of proteolytic events in apoptosis. Cell 134:679–691PubMedCentralPubMedCrossRefGoogle Scholar
  2. 2.
    Simon GM, Dix MM, Cravatt BF (2009) Comparative assessment of large-scale proteomic studies of apoptotic proteolysis. ACS Chem Biol 4:401–408PubMedCentralPubMedCrossRefGoogle Scholar
  3. 3.
    Cociorva D, Tabb DL, Yates JR (2006) Validation of tandem mass spectrometry database search results using DTASelect. Curr Protoc Bioinformatics 13.4.1–13.4.14, supplement 16, Johnwiley and SonsGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Melissa M. Dix
    • 1
  • Gabriel M. Simon
    • 2
  • Benjamin F. Cravatt
    • 1
  1. 1.Department of Chemical PhysiologyThe Scripps Research InstituteLa JollaUSA
  2. 2.Abide TherapeuticsLa JollaUSA

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