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Protocol for the production and reconstitution of VDAC1 for functional assays. | LitMetric

Protocol for the production and reconstitution of VDAC1 for functional assays.

STAR Protoc

Laboratoire d'Ingénierie des Systèmes Macromoléculaires, CNRS - Aix Marseille Université, 31 Chemin Joseph Aiguier, 13402 Marseille, France. Electronic address:

Published: September 2024

AI Article Synopsis

Article Abstract

The voltage-dependent anion channel (VDAC) is an abundant and multifunctional outer mitochondrial membrane protein, playing key roles in neurodegeneration, apoptosis, and mitochondrial membrane biogenesis. Here, we present a protocol to produce and reconstitute high yields of detergent-solubilized VDAC, expressed as inclusion bodies in E. coli. We describe steps for purification by affinity chromatography and refolding in lauryldimethylamine-N-oxide (LDAO). We then detail procedures for reconstituting VDAC into membrane vesicles to assay its channel and phospholipid scramblase activity via fluorescence-based assays. For complete details on the use and execution of this protocol, please refer to Bergdoll et al., Queralt-Martín et al., and Jahn et al..

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Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC11383923PMC
http://dx.doi.org/10.1016/j.xpro.2024.103240DOI Listing

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