title: Nitric oxide interacts with mitochondrial complex III producing antimycin-like effects.
abstract: The effect of NO between cytochromes b and c of the mitochondrial respiratory chain were studied using submitochondrial particles (SMP) from bovine heart and GSNO and SPER-NO as NO sources. Succinate-cytochrome c reductase (complex II-III) activity (222 ± 4 nmol/min. mg protein) was inhibited by 51% in the presence of 500 μM GSNO and by 48% in the presence of 30 μM SPER-NO, in both cases at ~1.25 μM NO. Neither GSNO nor SPER-NO were able to inhibit succinate-Q reductase activity (complex II; 220 ± 9 nmol/min. mg protein), showing that NO affects complex III. Complex II-III activity was decreased (36%) when SMP were incubated with l-arginine and mtNOS cofactors, indicating that this effect is also produced by endogenous NO. GSNO (500 μM) reduced cytochrome b562 by 71%, in an [O2] independent manner. Hyperbolic increases in O2(•-) (up to 1.3 ± 0.1 nmol/min. mg protein) and H2O2 (up to 0.64 ± 0.05 nmol/min. mg protein) productions were observed with a maximal effect at 500 μM GSNO. The O2(•-)/H2O2 ratio was 1.98 in accordance with the stoichiometry of the O2(•-) disproportionation. Moreover, H2O2 production was increased by 72-74% when heart coupled mitochondria were exposed to 500 μM GSNO or 30 μM SPER-NO. SMP incubated in the presence of succinate showed an EPR signal (g=1.99) compatible with a stable semiquinone. This EPR signal was increased not only by antimycin but also by GSNO and SPER-NO. These signals were not modified under N2 atmosphere, indicating that they are not a consequence to the effect of NOx species on complex III area. These results show that NO interacts with ubiquinone-cytochrome b area producing antimycin-like effects. This behaviour comprises the inhibition of electron transfer, the interruption of the oxidation of cytochromes b, and the enhancement of [UQH(•)]ss which, in turn, leads to an increase in O2(•-) and H2O2 mitochondrial production rates.
authors.affiliation: Institute of Biochemistry and Molecular Medicine, Physical Chemistry Division, School of Pharmacy and Biochemistry, University of Buenos Aires (IBIMOL, UBA-CONICET). Junín 956, C1113AAD Buenos Aires, Argentina. Electronic address: diglesias@ffyb.uba.ar.
authors.forename: Darío E
authors.initials: DE
authors.lastname: Iglesias
authors.affiliation: Institute of Biochemistry and Molecular Medicine, Physical Chemistry Division, School of Pharmacy and Biochemistry, University of Buenos Aires (IBIMOL, UBA-CONICET). Junín 956, C1113AAD Buenos Aires, Argentina.
authors.forename: Silvina S
authors.initials: SS
authors.lastname: Bombicino
authors.affiliation: Institute of Biochemistry and Molecular Medicine, Physical Chemistry Division, School of Pharmacy and Biochemistry, University of Buenos Aires (IBIMOL, UBA-CONICET). Junín 956, C1113AAD Buenos Aires, Argentina.
authors.forename: Laura B
authors.initials: LB
authors.lastname: Valdez
authors.affiliation: Institute of Biochemistry and Molecular Medicine, Physical Chemistry Division, School of Pharmacy and Biochemistry, University of Buenos Aires (IBIMOL, UBA-CONICET). Junín 956, C1113AAD Buenos Aires, Argentina.
authors.forename: Alberto
authors.initials: A
authors.lastname: Boveris
mesh_terms: D000818:Animals; D000968:Antimycin A / Q000031:analogs & derivatives / Q000378:metabolism; D002417:Cattle; D004578:Electron Spin Resonance Spectroscopy; D014450:Electron Transport Complex III / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000378:metabolism; D008929:Mitochondria, Heart / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D010084:Oxidation-Reduction; D051381:Rats; D013367:Submitochondrial Particles / Q000378:metabolism*
chemical_list: C032456:antimycin; D009569:Nitric Oxide; D000968:Antimycin A; D006861:Hydrogen Peroxide; D014450:Electron Transport Complex III
keywords: Cytochrome bc(1) complex; Electron paramagnetic resonance (EPR); Hydrogen peroxide; Nitric oxide; S-nitrosoglutathione (GSNO); Spermine-NONOate (SPER-NO); Superoxide anion; Ubiquinone-cytochrome b area; Ubisemiquinone