GST-Perfringolysin O production for the localization and quantification of membrane cholesterol in human and mouse brain and liver.

ABSTRACT

Abnormal cholesterol metabolism is linked to many neurodegenerative disorders. Here, we present a protocol for the production of a recombinant protein consisting of a Glutathione-S-Transferase tag fused with the Perfringolysin O (PFO). The GST-PFO tag enables analysis of the localization of cholesterol in subcellular membranes of human and mice brain and liver tissues. We have used this approach for samples from Niemann-Pick type C disease and non-alcoholic steatohepatitis models. The construct may also have applications for the diagnosis of cholesterol-accumulating disorders. For complete details on the use and execution of this protocol, please refer to Kwiatkowska et al. (2014).