REACTIVE OXYGEN SPECIES-MEDIATED INACTIVATION OF PYRUVATE-DEHYDROGENASE

Brain ischemia reperfusion causes increased formation of reactive oxyg en species (ROS). Activity of the mitochondrial enzyme pyruvate dehydr ogenase (PDH) has been shown to undergo a significant decrease followi ng reperfusion of the ischemic tissue. We have examined the effect of a superoxide radical-generating system (xanthine oxidase/hypoxanthine, XO/HX) on the activity of this enzyme. Incubation of PDH in the prese nce of XO/HX resulted in its inactivation. The degree of the inactivat ion was dependent on the amount of XO present, which correlated linear ly with the concentration of superoxide radical generated by this syst em. The activity of lactate dehydrogenase, an enzyme resistant to inac tivation by ischemia reperfusion, was not affected by this system. Sup eroxide dismutase partially prevented and catalase exerted a nearly co mplete protective effect against the inactivation of PDH. Deferoxamine was partially protective, The sulfhydryl protective reagents, dithiot hreitol and glutathione, prevented the inactivation of PDH, even thoug h to varying-degrees, which implicates sulfhydryl oxidation. A hydroxy l radical-generating system (hydrogen peroxide irradiated with ultravi olet radiation) effectively inactivated PDH. These results demonstrate that PDH is susceptible to damage and inactivation by ROS and point t o the involvement of Fenton chemistry and hydroxyl radicals formed thr ough it in PDH inactivation by XO/HX, A similar mechanism may be respo nsible for the PDH inactivation during ischemia/reperfusion. (C) 1996 Academic Press, Inc.