Modified Blue Native Gel Approach for Analysis of Respiratory Supercomplexes

Methods Mol Biol. 2021;2276:227-234. doi: 10.1007/978-1-0716-1266-8_17.

Abstract

In mitochondrial oxidative phosphorylation (Ox-Phos), individual electron transport chain complexes are thought to assemble into supramolecular entities termed supercomplexes (SCs). The technique of blue native (BN) gel electrophoresis has emerged as the method of choice for analyzing SCs. However, the process of sample extraction for BN gel analysis is somewhat tedious and introduces the possibility for experimental artifacts. Here we outline a streamlined method that eliminates a centrifugation step and provides a more representative sampling of a population of mitochondria on the final gel. Using this method, we show that SC composition does not appear to change dynamically with altered mitochondrial function.

Keywords: Blue-native; Clear-native; Mitochondria; Permeability transition pore; Respiration; Supercomplexes.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Electron Transport Chain Complex Proteins / analysis*
  • Electron Transport Chain Complex Proteins / metabolism
  • Heart / physiology*
  • Mice
  • Mitochondria, Heart / chemistry*
  • Mitochondria, Heart / metabolism
  • Mitochondrial Proteins / analysis*
  • Mitochondrial Proteins / metabolism
  • Multiprotein Complexes / analysis*
  • Multiprotein Complexes / metabolism
  • Myocardium / chemistry*
  • Myocardium / metabolism
  • Native Polyacrylamide Gel Electrophoresis / methods*
  • Oxidative Phosphorylation

Substances

  • Electron Transport Chain Complex Proteins
  • Mitochondrial Proteins
  • Multiprotein Complexes