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

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

Fingerprint

Immunoprecipitation
Spectrometry
Spectrum Analysis
Gels
Proteins
Native Polyacrylamide Gel Electrophoresis
Electrophoresis
Proteomics
Mass spectrometry
Mass Spectrometry
Chemical analysis

Keywords

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

Cite this

van der Spek, Sophie J.F. ; Smit, August B. ; Pandya, Nikhil J. / Integrated immunoprecipitation : Blue native gel electrophoresis—mass spectrometry for the identification of protein subcomplexes. Neuroproteomics. editor / Ka Wan Li. 2nd. ed. Humana Press Inc., 2019. pp. 85-94 (Neuromethods).
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Integrated immunoprecipitation : Blue native gel electrophoresis—mass spectrometry for the identification of protein subcomplexes. / van der Spek, Sophie J.F.; Smit, August B.; Pandya, Nikhil J.

Neuroproteomics. ed. / Ka Wan Li. 2nd. ed. Humana Press Inc., 2019. p. 85-94 (Neuromethods; Vol. 146).

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

TY - CHAP

T1 - Integrated immunoprecipitation

T2 - Blue native gel electrophoresis—mass spectrometry for the identification of protein subcomplexes

AU - van der Spek, Sophie J.F.

AU - Smit, August B.

AU - Pandya, Nikhil J.

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AB - 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.

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KW - Proteomics

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