Kinetic mechanism of nitric oxide-inhibited mitochondrial complex III. Effect on ubisemiquinone and superoxide formation

IGLESIAS DE; BOMBICINO SS; BOVERIS A; VALDEZ LB

Ferrara

Congreso; SFRR-E 2019 Annual Meeting. Redox homeostasis: from signaling to damage?; 2019

Society for Free Radical Research-Europe

We have shown that NO inhibits complex III, increasing [UQH●]ss, [cyt. b2+], and O2●- and H2O2 production rates. This work investigated the interaction of NO with complex III using a complex III enriched fraction -isolated from bovine heart- and NO solution (obtained by bubbling NO; 10% in N2), in order to characterize the inhibition of electron transfer and the consequences on O2●- production, and to postulate a kinetic mechanism that explains the experimental results. Complex III exposed to NO showed a hyperbolic inhibition of decylubiquinol:cytochrome c reductase activity (IC50=225 nM) and the characteristic peak of cyt. bH2+, suggesting the interaction of NO with the hemo b562. According to [UQH●]ss enhancement detected by EPR, H2O2 production rate was augmented (55%) in coupled mitochondria exposed to 1 M NO. These results can be explained by a kinetic model that considers the interaction of NO with cyt. bH and, consequently, the accumulation of cyt. bL2+, due to the blockage of electron transfer between cytochromes b. In this scenario, the formation of UQH● is mainly driven by reverse electron flow from cyt. bL2+ to ubiquinone, explaining the increase of O2●- production. Therefore, NO physiologically generated can act as the non physiological complex III inhibitor antimycin.