Integrated immunoprecipitation: Blue native gel electrophoresis—mass spectrometry for the identification of protein subcomplexes

Sophie J.F. van der Spek, August B. Smit, Nikhil J. Pandya*

*Corresponding author for this work

Research output: Chapter in Book / Report / Conference proceedingChapterAcademicpeer-review

Abstract

Proteins typically act as components of poly-protein complexes, which may vary in composition and function, and may depend on subcellular localization and cellular state. To understand their biology, there is a need to identify the constituents in these various protein complexes. Conventional immunoprecipitation combined with proteomics analysis is nowadays routinely used to identify multiple interactors of a bait protein; however, it is unable to distinguish protein subcomplexes from one another. Blue native polyacrylamide gel electrophoresis is appropriate to separate individual protein complexes. Here, we describe a protocol that combines the specificity of immunoprecipitation, the size separating capacity of Blue native polyacrylamide gel electrophoresis, and subsequent mass spectrometry to delineate the protein constituents in subcomplexes of a targeted protein.

Original languageEnglish
Title of host publicationNeuroproteomics
EditorsKa Wan Li
PublisherHumana Press Inc.
Pages85-94
Number of pages10
Edition2nd
ISBN (Electronic)9781493996629
ISBN (Print)9781493996612
DOIs
Publication statusPublished - 2019

Publication series

NameNeuromethods
Volume146
ISSN (Print)0893-2336
ISSN (Electronic)1940-6045

Keywords

  • Blue native polyacrylamide gel electrophoresis
  • Immunoprecipitation
  • Mass spectrometry
  • Protein complexes
  • Proteomics

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