Protein S-glutathionylation alters superoxide/hydrogen peroxide emission from pyruvate dehydrogenase complex

Publication date: May 2017
Source:Free Radical Biology and Medicine, Volume 106
Author(s): Marisa O'Brien, Julia Chalker, Liam Slade, Danielle Gardiner, Ryan J. Mailloux
Pyruvate dehydrogenase (Pdh) is a vital source of reactive oxygen species (ROS) in several different tissues. Pdh has also been suggested to serve as a mitochondrial redox sensor. Here, we report that O2^•-/ H2O2 emission from pyruvate dehydrogenase (Pdh) is altered by S-glutathionylation. Glutathione disulfide (GSSG) amplified O2^•-/ H2O2 production by purified Pdh during reverse electron transfer (RET) from NADH. Thiol oxidoreductase glutaredoxin-2 (Grx2) reversed these effects confirming that Pdh is a target for S-glutathionylation. S-glutathionylation had the opposite effect during forward electron transfer (FET) from pyruvate to NAD⁺ lowering O2^•-/ H2O2 production. Immunoblotting for protein glutathione mixed disulfides (PSSG) following diamide treatment confirmed that purified Pdh can be S-glutathionylated. Similar observations were made with mouse liver mitochondria. S-glutathionylation catalysts diamide and disulfiram significantly reduced pyruvate or 2-oxoglutarate driven O2^•-/ H2O2 production in liver mitochondria, results that were confirmed using various Pdh, 2-oxoglutarate dehydrogenase (Ogdh), and respiratory chain inhibitors. Immunoprecipitation of Pdh and Ogdh confirmed that either protein can be S-glutathionylated by diamide and disulfiram. Collectively, our results demonstrate that the S -glutathionylation of Pdh alters the amount of ROS formed by the enzyme complex. We also confirmed that Ogdh is controlled in a similar manner. Taken together, our results indicate that the redox sensing and ROS forming properties of Pdh and Ogdh are linked to S-glutathionylation.



http://ift.tt/2mIkGx3