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Mass Spectrometry of Mitochondrial Membrane Protein Complexes

  • Luc Negroni
  • Michel Zivy
  • Claire LemaireEmail author
Protocol
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Part of the Methods in Molecular Biology book series (MIMB, volume 1635)

Abstract

The ATP production (oxidative phosphorylation) involves five complexes embedded in the inner membrane of mitochondria. The yeast Saccharomyces cerevisiae is mainly used as a model for the study of oxidative phosphorylation; mutants are easy to produce and are still viable due to their ability to grow using the fermentation pathway. Here, we present a process for analyzing mitochondrial respiratory complexes using native electrophoresis (BN-PAGE) coupled to LC-MS/MS. BN-PAGE (1) permits the separation of functional respiratory complexes, thus allowing in-gel activity detection of most of the respiratory complexes and (2) provides convenient samples for bottom-up proteomics. Combining BN-PAGE and LC-MS/MS leads to the identification of the subunit composition of membrane complexes and offers the possibility of highlighting potential interacting proteins.

Key words

S. cerevisiae Respiratory complexes OXPHOS BN-PAGE Mass spectrometry 

Notes

Acknowledgments

We gratefully acknowledge Alain Guillot for updates made to the technical points and for critical reading of the manuscript.

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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  1. 1.CNRS-UMR5248Université de BordeauxBordeauxFrance
  2. 2.IGBMC, CNRS-UMR 7104IllkirchFrance
  3. 3.CNRS, PAPPSOUMR Génétique Quantitative et Evolution – Le MoulonGif-sur-YvetteFrance
  4. 4.CNRS-UMR9198, CEA-IBITECSUniversité Paris-Sud, I2BCGif-sur-YvetteFrance

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