Assessment of Mitochondrial Protein Glutathionylation as Signaling for CO Pathway
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Protein glutathionylation is a posttranslational process that regulates protein function in response to redox cellular changes. Furthermore, carbon monoxide-induced cellular pathways involve reactive oxygen species (ROS) signaling and mitochondrial protein glutathionylation. Herein, it is described a technique to assess mitochondrial glutathionylation due to low concentrations of CO exposure. Mitochondria are isolated from cell culture or tissue, followed by an immunoprecipitation assay, which allows the capture of any glutathionylated mitochondrial protein using a specific antibody coupled to a solid matrix that binds to glutathione antigen. The precipitated protein is further identified and quantified by immunoblotting analysis.
Key wordsGlutathionylation Carbon monoxide Mitochondria Glutathione Immunoprecipitation
This work was supported by the Portuguese Fundação para a Ciência e a Tecnologia (FCT-ANR/NEU-NMC/0022/2012) and the Portuguese Fundação para a Ciência e a Tecnologia for ASA’s SFRH/BD/78440/2011 fellowship.