Selective inhibition by 2-heptyl-4-hydroxyquinoline N-oxide of certain oxidation-reduction reactions

Abstract

The streptomycin antagonist, 2-heptyl-4-hydroxyquinoline N-oxide, inhibited the oxidation of dihydro-nicotinamide-adenine dinucleotide (NADH2) by hydrogen peroxide or methyl hydrogen peroxide, catalyzed by a NADH2 peroxidase, isolated in purified form from Bacillus subtilis. This enzyme is a flavoprotein which, in its purified form, required flavin-adenine dinucleotide (FAD) for activity. The inhibition by 2-heptyl-4-hydroxyquinoline N-oxide was only observed in enzyme preparations which showed requirement for FAD. The inhibition of NADH2 peroxidase by 2-heptyl-4-hydroxy-quinoline N-oxide was non-competitive with respect to NADH2, peroxide or added FAD. Experiments in which the reduction of FAD by NADH2 catalyzed by NADH2 peroxidase was measured indicated that, in the presence of enzyme, 2-heptyl-4-hydroxyquinoline N-oxide altered the oxidation-reduction properties of FAD. Experiments with non-enzymic model systems indicated that 2-heptyl-4-hydroxyquinoline N-oxide also altered the oxidation-reduction properties of free flavins and could enhance the formation of a semiquinoid complex formed by flavin mononucleotide in the presence of tryptophan. Experiments with model systems in which the oxidation of reduced 2,6-dichlorophenol-indophenol is catalyzed by cytochrome c, haematin, horse-radish peroxidase or a bacterial cytochrome from Pseudomonas, showed that 2-heptyl-4-hydroxyquinoline N-oxide inhibited the reaction in the presence of the first two, but not with the other two. The inhibition appeared to be localized on the oxidation side of the haem compound. Difference spectra of mammalian cytochrome c and cytochrome c in the presence of 2-heptyl-4-hydroxyquinoline N-oxide showed that some of the absorption of cytochrome c disappeared in the presence of 2-heptyl-4-hydroxyquinoline N-oxide. The effect, though proportional to concentrations of 2-heptyl-4-hydroxyquinoline N-oxide, was small and suggested either a very low affinity of cytochrome c for 2-heptyl-4-hydroxyquinoline N-oxide or, alternatively, the formation of a complex between 2-heptyl-4-hydroxyquinoline N-oxide and cytochrome c with a spectrum similar to that of cytochrome c but with different extinction coefficients.