ubiquinol and quinone

Topics: Amino Acid Sequence; Animals; Benzoquinones; Binding Sites; Cattle; Cell Membrane; Electron Transport Complex III; Electrons; Mitochondria; Models, Biological; Models, Molecular; Molecular Sequence Data; Mutation; Myocardium; Oxidation-Reduction; Protein Binding; Protons; Ubiquinone

Since available structures of native bc(1) complexes show a vacant Q(o)-site, occupancy by substrate and product must be investigated by kinetic and spectroscopic approaches. In this brief review, we discuss recent advances using these approaches that throw new light on the mechanism. The rate-limiting reaction is the first electron transfer after formation of the enzyme-substrate complex at the Q(o)-site. This is formed by binding of both ubiquinol (QH(2)) and the dissociated oxidized iron-sulfur protein (ISP(ox)). A binding constant of approximately 14 can be estimated from the displacement of E(m) or pK for quinone or ISP(ox), respectively. The binding likely involves a hydrogen bond, through which a proton-coupled electron transfer occurs. An enzyme-product complex is also formed at the Q(o)-site, in which ubiquinone (Q) hydrogen bonds with the reduced ISP (ISPH). The complex has been characterized in ESEEM experiments, which detect a histidine ligand, likely His-161 of ISP (in mitochondrial numbering), with a configuration similar to that in the complex of ISPH with stigmatellin. This special configuration is lost on binding of myxothiazol. Formation of the H-bond has been explored through the redox dependence of cytochrome c oxidation. We confirm previous reports of a decrease in E(m) of ISP on addition of myxothiazol, and show that this change can be detected kinetically. We suggest that the myxothiazol-induced change reflects loss of the interaction of ISPH with Q, and that the change in E(m) reflects a binding constant of approximately 4. We discuss previous data in the light of this new hypothesis, and suggest that the native structure might involve a less than optimal configuration that lowers the binding energy of complexes formed at the Q(o)-site so as to favor dissociation. We also discuss recent results from studies of the bypass reactions at the site, which lead to superoxide (SO) production under aerobic conditions, and provide additional information about intermediate states.

Topics: Benzoquinones; Binding Sites; Electron Transport Complex III; Iron-Sulfur Proteins; Kinetics; Methacrylates; Oxidation-Reduction; Thermodynamics; Thiazoles; Ubiquinone

We compared the influence of different adenine and guanine nucleotides on the free fatty acid-induced uncoupling protein (UCP) activity in non-phosphorylating Acanthamoeba castellanii mitochondria when the membranous ubiquinone (Q) redox state was varied. The purine nucleotides exhibit an inhibitory effect in the following descending order: GTP>ATP>GDP>ADP≫GMP>AMP. The efficiency of guanine and adenine nucleotides to inhibit UCP-sustained uncoupling in A. castellanii mitochondria depends on the Q redox state. Inhibition by purine nucleotides can be increased with decreasing Q reduction level (thereby ubiquinol, QH₂ concentration) even with nucleoside monophosphates that are very weak inhibitors at the initial respiration. On the other hand, the inhibition can be alleviated with increasing Q reduction level (thereby QH₂ concentration). The most important finding was that ubiquinol (QH₂) but not oxidised Q functions as a negative regulator of UCP inhibition by purine nucleotides. For a given concentration of QH₂, the linoleic acid-induced GTP-inhibited H(+) leak was the same for two types of A. castellanii mitochondria that differ in the endogenous Q content. When availability of the inhibitor (GTP) or the negative inhibition modulator (QH₂) was changed, a competitive influence on the UCP activity was observed. QH₂ decreases the affinity of UCP for GTP and, vice versa, GTP decreases the affinity of UCP for QH₂. These results describe the kinetic mechanism of regulation of UCP affinity for purine nucleotides by endogenous QH₂ in the mitochondria of a unicellular eukaryote.

Topics: Acanthamoeba castellanii; Adenine Nucleotides; Benzoquinones; Fatty Acids, Nonesterified; Guanine Nucleotides; Homeostasis; Ion Channels; Membrane Potentials; Mitochondria; Mitochondrial Proteins; Oxidation-Reduction; Oxygen Consumption; Purine Nucleotides; Ribonucleotides; Ubiquinone; Uncoupling Protein 1

A non-photosynthetic mutant (Ps-) of Rhodopseudomonas capsulata, designated R126, was analyzed for a defect in the cyclic electron transfer system. Compared to a Ps+ strain MR126, the mutant was shown to have a full complement of electron transfer components (reaction centers, ubiquinone-10, cytochromes b, c1, and c2, the Rieske 2-iron, 2-sulfur (Rieske FeS) center, and the antimycin-sensitive semiquinone). Functionally, mutant R126 failed to catalyze complete cytochrome c1 + c2 re-reduction or cytochrome b reduction following a short (10 microseconds) flash of actinic light. Evidence (from flash-induced carotenoid band shift) was characteristic of inhibition of electron transfer proximal to cytochrome c1 of the ubiquinol-cytochrome c2 oxidoreductase. Three lines of evidence indicate that the lesion of R126 disrupts electron transfer from quinol to Rieske FeS: 1) the degree of cytochrome c1 + c2 re-reduction following a flash is indicative of electron transfer from Rieske FeS to cytochrome c1 + c2 without redox equilibration with an additional electron from a quinol; 2) inhibitors that act at the Qz site and raise the Rieske FeS midpoint redox potential (Em), namely 5-undecyl-6-hydroxy-4,7-dioxobenzothiazole or 3-alkyl-2-hydroxy-1,4-napthoquinone, have no effect on cytochrome c1 + c2 oxidation in R126; 3) the Rieske FeS center, although it exhibits normal redox behavior, is unable to report the redox state of the quinone pool, as metered by its EPR line shape properties. Flash-induced proton binding in R126 is indicative of normal functional primary (QA) and secondary (QB) electron acceptor activity of the photosynthetic reaction center. The Qc functional site of cytochrome bc1 is intact in R126 as measured by the existence of antimycin-sensitive, flash-induced cytochrome b reduction.

Topics: Antimycin A; Benzoquinones; Cytochrome c Group; Electron Spin Resonance Spectroscopy; Electron Transport; Electron Transport Complex III; Methacrylates; Multienzyme Complexes; Mutation; Oxidation-Reduction; Photolysis; Quinone Reductases; Quinones; Rhodopseudomonas; Thiazoles; Ubiquinone