Reactions of peroxynitrite in the mitochondrial matrix.

VALDEZ, L.; ALVAREZ, S.; LORES ARNAIZ, S.; SCHOPFER, F.; PODEROSO, J.J. ; BOVERIS, A.

FREE RADICAL BIOLOGY AND MEDICINE

ELSEVIER SCIENCE INC

Lugar: Amsterdam; Año: 2000 vol. 29 p. 349 - 356

0891-5849

Superoxide radical (O22 2) and nitric oxide (NO) produced at the mitochondrial inner membrane react to form peroxynitrite (ONOO2) in the mitochondrial matrix. Intramitochondrial ONOO2 effectively reacts with a few biomolecules according to reaction constants and intramitochondrial concentrations. The second-order reaction constants (in M21 s21) of ONOO2 with NADH (233 6 27), ubiquinol-0 (485 6 54) and GSH (183 6 12) were determined fluorometrically by a simple competition assay of product formation. The oxidation of the components of the mitochondrial matrix by ONOO2 was also followed in the presence of CO2, to assess the reactivity of the nitrosoperoxocarboxylate adduct (ONOOCO2) and nitric oxide (NO) produced at the mitochondrial inner membrane react to form peroxynitrite (ONOO2) in the mitochondrial matrix. Intramitochondrial ONOO2 effectively reacts with a few biomolecules according to reaction constants and intramitochondrial concentrations. The second-order reaction constants (in M21 s21) of ONOO2 with NADH (233 6 27), ubiquinol-0 (485 6 54) and GSH (183 6 12) were determined fluorometrically by a simple competition assay of product formation. The oxidation of the components of the mitochondrial matrix by ONOO2 was also followed in the presence of CO2, to assess the reactivity of the nitrosoperoxocarboxylate adduct (ONOOCO22) in the mitochondrial matrix. Intramitochondrial ONOO2 effectively reacts with a few biomolecules according to reaction constants and intramitochondrial concentrations. The second-order reaction constants (in M21 s21) of ONOO2 with NADH (233 6 27), ubiquinol-0 (485 6 54) and GSH (183 6 12) were determined fluorometrically by a simple competition assay of product formation. The oxidation of the components of the mitochondrial matrix by ONOO2 was also followed in the presence of CO2, to assess the reactivity of the nitrosoperoxocarboxylate adduct (ONOOCO221 s21) of ONOO2 with NADH (233 6 27), ubiquinol-0 (485 6 54) and GSH (183 6 12) were determined fluorometrically by a simple competition assay of product formation. The oxidation of the components of the mitochondrial matrix by ONOO2 was also followed in the presence of CO2, to assess the reactivity of the nitrosoperoxocarboxylate adduct (ONOOCO22 was also followed in the presence of CO2, to assess the reactivity of the nitrosoperoxocarboxylate adduct (ONOOCO22 2) towards the same reductants. The ratio of product formation was about similar both in the presence of 2.5 mM CO2 and in air-equilibrated conditions. Liver submitochondrial particles supplemented with 0.25–2 mM ONOO2 showed a O2) towards the same reductants. The ratio of product formation was about similar both in the presence of 2.5 mM CO2 and in air-equilibrated conditions. Liver submitochondrial particles supplemented with 0.25–2 mM ONOO2 showed a O22 and in air-equilibrated conditions. Liver submitochondrial particles supplemented with 0.25–2 mM ONOO2 showed a O2mM ONOO2 showed a O2 2 production that indicated ubisemiquinone formation and autooxidation. The nitration of mitochondrial proteins produced after addition of 200 mM ONOO2 was observed by Western blot analysis. Protein nitration was prevented by the addition of 50–200 mM ubiquinol-0 or GSH. An intramitochondrial steady state concentration of abouproduction that indicated ubisemiquinone formation and autooxidation. The nitration of mitochondrial proteins produced after addition of 200 mM ONOO2 was observed by Western blot analysis. Protein nitration was prevented by the addition of 50–200 mM ubiquinol-0 or GSH. An intramitochondrial steady state concentration of aboumM ONOO2 was observed by Western blot analysis. Protein nitration was prevented by the addition of 50–200 mM ubiquinol-0 or GSH. An intramitochondrial steady state concentration of aboumM ubiquinol-0 or GSH. An intramitochondrial steady state concentration of abou