Interaction of NO with mitochondrial Complex III: functional consequences and mechanism

Previous results from our laboratory showed that NO inhibits complex III producing antimycin-like effects regarding to the enhancements of [UQH•]ss, [cyt. b2+], and O2•- and H2O2 productions. In this work, the effects of NO solution (obtained by bubbling NO gas; 10% in N2), on complex III enriched fraction -isolated from bovine heart- were studied. Complex III activity was hyperbolic inhibited by NO (IC50=225 nM). The absorbance spectra of complex III exposed to NO showed the characteristic peak of cyt. bH2+ (562 nm). In coupled mitochondria, 1 µM NO inhibited succinate-sustained state 3 O2 consumption (50%). Addition of HbO2 recovered O2 consumption by 90%; the remaining 10% was insensitive to NO scavenging, suggesting the blockage of complex III by NO. Moreover, mitochondria exposed to NO showed a less change in ΔΨ (30%) in the transition from state 4 to state 3 respiration, lowering ATP synthesis capacity. According to [UQH•]ss enhancement detected by EPR, H2O2 production rate was augmented (55%). Altogether, these results could be explained through a kinetic model which considers the interaction of NO with cyt. bH and, consequently, the inhibition of electron transfer between cytochromes b, the formation of UQH•, and the increase of O2•- and H2O2 production rates.