chiniofon and funiculosin-(anthraquinone)

Topics: Anthraquinones; Antifungal Agents; Antimycin A; Cytochrome b Group; Diuron; Frameshift Mutation; Hydroxyquinolines; Mitochondria; Point Mutation; Protein Conformation; Protein Structure, Tertiary; Recombinant Proteins; Saccharomyces cerevisiae; Spectrophotometry

The effects of pH and inhibitors on the spectra and redox properties of the haems b of the bc1 complex of beef heart submitochondrial particles were investigated. The major findings were: (1) both haems have a weakly redox-linked protonatable group with pKox and pKred of around 6 and 8; (2) at pH values above 7, haem bH becomes heterogeneous in its redox behaviour. This heterogeneity is removed by the Qi site inhibitors antimycin A, funiculosin and HQNO, but not by the Qo site inhibitors myxothiazol or stigmatellin; (3) of all inhibitors tested only funiculosin had a large effect on the Em/pH profile of either haem b. In all cases where definite effects were found, the haem most affected was that thought to be closest to the site of inhibitor binding; (4) spectral shifts of haem groups caused by inhibitor binding were usually, but not always, of the haem group closest to the binding site; (5) titrations with succinate/fumarate were in reasonable agreement with redox-mediated data provided that strict anaerobiosis was maintained. Apparent large shifts of haem midpoint potentials with antimycin A and myxothiazol could be produced in aerobic succinate/fumarate titrations in the presence of cyanide, as already reported in the literature, but these were artefactual; (6) the heterogeneous haem bH titration behaviour can be simulated with a model similar to that proposed by Salerno et al. (J. Biol. Chem. (1989) 264, 15398-15403) in which there is redox interaction between haem bH and ubiquinone species bound at the Qi site. Simulations closely fit both the haem bH data and known semiquinone data only if it is assumed that semiquinone bound to oxidised haem bH is EPR-silent.

Topics: Animals; Anthraquinones; Antimycin A; Cattle; Electron Spin Resonance Spectroscopy; Electron Transport Complex III; Fumarates; Heme; Hydrogen-Ion Concentration; Hydroxyquinolines; Mitochondria, Heart; Oxidation-Reduction; Protons; Submitochondrial Particles; Succinates; Succinic Acid

O2-. generation by the succinate oxidase segment of the respiratory chain of mitochondria and submitochondrial particles from hepatoma 22a and hepatoma Zajdela has been studied with the use of the Tiron method. In the presence of succinate, superoxide generation is induced by antimycin, 2-n-4-hydroxyquinoline N-oxide or funiculosin, and is inhibited by mucidin, myxothiazol or cyanide. The rate of O2-. generation in the antimycin-inhibited state is maximal at the [succinate]/[fumarate] ratio of 1:10 and diminishes at more positive and more negative redox potentials. These characteristics of O2-.-generation are the same as observed earlier in submitochondrial particles from normal tissues. Accordingly, the mechanism of superoxide production is suggested to be the same in tumor and normal mitochondria, namely, autoxidation of the unstable ubisemiquinone in the ubiquinol-oxidizing centre o of cytochrome bc1 complex. With respect to the rate of O2-. generation, the hepatoma mitochondrial membranes are approximately twice as active as bovine heart submitochondrial particles and exceed those from rat liver by more than one order of magnitude.

Topics: 1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt; Animals; Anthraquinones; Antimycin A; Benzoquinones; Electron Spin Resonance Spectroscopy; Electron Transport; Free Radicals; Hydroxyquinolines; Kinetics; Liver Neoplasms, Experimental; Male; Mice; Mice, Inbred C3H; Mitochondria; Oxidation-Reduction; Oxidoreductases; Quinones; Rats; Submitochondrial Particles; Succinates; Succinic Acid; Superoxides

Changes are described that are brought about by antimycin, NoHOQnO, funiculosin, myxothiazol and mucidin in the alpha-, beta- and gamma-absorption bands of reduced and oxidized cytochromes b in the isolated complex bc1 form beef heart mitochondria. The inhibitors can be divided into 2 groups. Antimycin, funiculosin and NoHOQnO are likely to shift the spectrum of b-562 and compete for specific binding with complex bc1, with each other but not with myxothiazol and mucidin. The spectral effects of the latter two inhibitors are more difficult to interpret and may involve contributions not only from b-562 but from b-566 as well. The existence of 2 independent inhibitor binding-sites in the complex bc1 corroborates the Q-cycle hypothesis.

Topics: Alkenes; Animals; Anthraquinones; Antimycin A; Cattle; Cytochrome b Group; Electron Transport; Electron Transport Complex III; Fatty Acids, Unsaturated; Hydroxyquinolines; Methacrylates; Mitochondria, Heart; Multienzyme Complexes; NADH, NADPH Oxidoreductases; Quinone Reductases; Spectrophotometry; Strobilurins; Thiazoles

Bovine-heart Complex III can catalyze the reduction of spinach plastocyanin by a decyl analog of ubiquinol-2 at a rate comparable with the rate of plastocyanin reduction by plastoquinol as catalyzed by the cytochrome b6-f complex purified from spinach leaves. This plastocyanin reduction as catalyzed by Complex III was almost completely inhibited by myxothiazol at stoichiometric concentrations, partially inhibited by UHDBT (5-n-undecyl-6-hydroxy-4,7-dioxobenzothiazole) and funiculosin, and was relatively insensitive to antimycin and HQNO (2-n-heptyl-4-hydroxyquinoline-N-oxide). Cytochrome c reduction as catalyzed by Complex III displayed a residual, inhibitor-insensitive rate of 5% of the uninhibited rate for each of the three inhibitors, antimycin, myxothiazol, and UHDBT. However, the residual rate that was insensitive to each of the inhibitors added singly was inhibited further by addition of the remaining two inhibitors. From these results it is concluded that plastocyanin reduction involves an electron-transfer pathway through Complex III that is distinct from the pathway utilized for reduction of cytochrome c.

Topics: Animals; Anthraquinones; Antimycin A; Cattle; Drug Resistance; Electron Transport; Electron Transport Complex III; Hydrogen-Ion Concentration; Hydroxyquinolines; Methacrylates; Models, Chemical; Multienzyme Complexes; Myocardium; NADH, NADPH Oxidoreductases; Oxidation-Reduction; Plant Proteins; Plastocyanin; Quinone Reductases; Thiazoles

Antimycin, 2-nonyl-4-hydroxyquinoline N-oxide and funiculosin induce O.2(-) generation by submitochondrial particles oxidizing succinate, whereas KCN, mucidin, myxothiazol or 2,3-dimercaptopropanol inhibit O.2(-) generation. Thenoyltrifluoroacetone does not induce superoxide production by itself but slightly stimulates the reaction initiated by antimycin. The results indicate that auto-oxidation of unstable ubisemiquinone formed in centre o of the Q-cycle generates most of the O.2(-) radicals in the cytochrome bc1-site of the mitochondrial respiratory chain.

Topics: Alkenes; Animals; Anthraquinones; Antimycin A; Cattle; Dimercaprol; Electron Transport Complex III; Fatty Acids, Unsaturated; Hydroxyquinolines; Methacrylates; Mitochondria, Heart; Multienzyme Complexes; NADH, NADPH Oxidoreductases; Oxygen; Quinone Reductases; Strobilurins; Superoxides; Thenoyltrifluoroacetone; Thiazoles

Anaerobic potentiometric titrations of b cytochromes have been carried out in beef heart submitochondrial particles in the presence of several specific inhibitors of electron transfer through the b-c1-site of the respiratory chain. Whereas antimycin shows no significant effect on the titration curve of cytochrome b-562, NoHOQnO is found to shift the Em of b-562 by 20-30 mV to the positive. Funiculosin raises the Em of b-562 by greater than 100 mV and also appears to bring about a minor shift of b-566 midpoint potential. In the presence of myxothiazol, both b cytochromes titrate with Em values 15-30 mV more positive than in the control.

Topics: Animals; Anthraquinones; Antimycin A; Binding Sites; Cattle; Cytochromes; Electron Transport Complex III; Hydroxyquinolines; Methacrylates; Mitochondria, Heart; Multienzyme Complexes; NADH, NADPH Oxidoreductases; Oxidation-Reduction; Potentiometry; Quinone Reductases; Thiazoles