Protocol to characterize mitochondrial supercomplexes from mouse tissues by combining BN-PAGE and MS-based proteomics

Roger Moreno-Justicia; Alba Gonzalez-Franquesa; Ben Stocks; Atul S Deshmukh

doi:10.1016/j.xpro.2022.101135

Protocol to characterize mitochondrial supercomplexes from mouse tissues by combining BN-PAGE and MS-based proteomics

STAR Protoc. 2022 Jan 28;3(1):101135. doi: 10.1016/j.xpro.2022.101135. eCollection 2022 Mar 18.

Authors

Roger Moreno-Justicia¹, Alba Gonzalez-Franquesa¹, Ben Stocks¹, Atul S Deshmukh¹

Affiliation

¹ Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, Copenhagen 2200, Denmark.

Abstract

The assembly of mitochondrial respiratory complexes into supercomplexes has significant implications for mitochondrial function. This protocol details mitochondrial isolation from mouse tissues and the use of blue native gel electrophoresis (BN-PAGE) to separate pre-identified mitochondrial supercomplexes into different gel bands. We then describe the excision of the individual bands, followed by in-gel protein digestion and peptide desalting for mass spectrometry (MS)-based proteomics. This protocol provides a time-efficient measurement of the abundance and distribution of proteins within known supercomplexes. For complete details on the use and execution of this profile, please refer to Gonzalez-Franquesa et al. (2021).

Keywords: Mass Spectrometry; Metabolism; Protein Biochemistry; Proteomics.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Electrophoresis
Mass Spectrometry
Mice
Mitochondria* / chemistry
Proteomics* / methods