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Query: UNIPROT:O95477 (membrane-bound)
 29,236 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

It was shown that the membrane-bound complex I is fully inactive in the absence of NADH during the reverse electron transfer from succinate to NAD+. The enzyme activation is attained by preincubation of submitochondrial particles with low concentrations of NADH; the activating effect persists after a complete oxidation of the latter during long-term (several hours) aerobic incubation. The experimental results suggest that complex I contains a redox component, whose reduction by NADH and aerobic oxidation are not involved in the overall catalytic reaction. An experimental scheme is proposed, according to which the key role of such a component is ascribed to the tightly bound ubiquinone; the activation and inactivation of the enzyme are due to a slow reversible redox conversion (ubiquinone in equilibrium ubisemiquinone), whereas the catalytic act involves a rapid reversible conversion (ubisemiquinone in equilibrium ubiquinol). It was demonstrated that the "redox" mechanism of the inactivation-activation reaction determines the strong dependence of activity of the reverse electron transfer on the mode of preparation of submitochondrial particles. The coupling properties of the submitochondrial particulate membrane and the activities of enzymes involved in the reverse electron transfer are stable at room temperature for over 14 hours.
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PMID:[Hysteresis behavior of complex I in delta mu H+-dependent reduction of NAD+ succinate]. 249 1

Acinetobacter calcoaceticus is known to contain soluble and membrane-bound quinoprotein D-glucose dehydrogenases while other oxidative bacteria such as Pseudomonas or Gluconobacter contain only membrane-bound enzyme. The two different forms were believed to be the same enzyme or interconvertible. Present results show that the two different forms of glucose dehydrogenase are distinct from each other in their enzymatic and immunological properties as well as in their molecular size. The soluble and membrane-bound glucose dehydrogenases were separated after French press-disruption by repeated ultracentrifugation, and then purified to nearly homogeneous state. The soluble enzyme was a polypeptide of 55 Kdaltons, while the membrane-bound enzyme was a polypeptide of 83 Kdaltons which is mainly monomeric in detergent solution. Both enzymes showed different enzymatic properties including substrate specificity, optimum pH, kinetics for glucose, and reactivity for ubiquinone-homologues. Furthermore, the two enzymes could be distinguished immunochemically; the membrane-bound enzyme is cross-reactive with an antibody raised against membrane-bound enzyme purified from Pseudomonas but not with antibody elicited against the soluble enzyme, while the soluble enzyme is not cross-reactive with the antibody of membrane-bound enzyme. Data also suggest that the membrane-bound enzyme functions by linking to the respiratory chain via ubiquinone though the function of the soluble enzyme remains unclear.
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PMID:Quinoprotein D-glucose dehydrogenases in Acinetobacter calcoaceticus LMD 79.41: purification and characterization of the membrane-bound enzyme distinct from the soluble enzyme. 254 65

Acinetobacter calcoaceticus is known to contain soluble and membrane-bound quinoprotein D-glucose dehydrogenases, while other oxidative bacteria contain the membrane-bound enzyme exclusively. The two forms of glucose dehydrogenase were believed to be the same enzyme or interconvertible forms. Previously, Matsushita et al. [(1988) FEMS Microbiol. Lett 55, 53-58] showed that the two enzymes are different with respect to enzymatic and immunological properties, as well as molecular weight. In the present study, we purified both enzymes and compared their kinetics, reactivity with ubiquinone homologues, and immunological properties in detail. The purified membrane-bound enzyme had a molecular weight of 83,000, while the soluble form was 55,000. The purified enzymes exhibited totally different enzymatic properties, particularly with respect to reactivity toward ubiquinone homologues. The soluble enzyme reacted with short-chain homologues only, whereas the membrane-bound enzyme reacted with long-chain homologues including ubiquinone 9, the native ubiquinone of the A. calcoaceticus. Furthermore, the two enzymes were distinguished immunochemically; the membrane-bound enzyme did not cross-react with antibody raised against the soluble enzyme, nor did the soluble enzyme cross-react with antibody against the membrane-bound enzyme. Thus, each glucose dehydrogenase is a molecularly distinct entity, and the membrane-bound enzyme only is coupled to the respiratory chain via ubiquinone.
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PMID:Quinoprotein D-glucose dehydrogenase of the Acinetobacter calcoaceticus respiratory chain: membrane-bound and soluble forms are different molecular species. 255 69

Membrane fluidity plays an important role in cellular functions. Membrane proteins are mobile in the lipid fluid environment; lateral diffusion of membrane proteins is slower than expected by theory, due to both the effect of protein crowding in the membrane and to constraints from the aqueous matrix. A major aspect of diffusion is in macromolecular associations: reduction of dimensionality for membrane diffusion facilitates collisional encounters, as those concerned with receptor-mediated signal transduction and with electron transfer chains. In mitochondrial electron transfer, diffusional control is prevented by the excess of collisional encounters between fast-diffusing ubiquinone and the respiratory complexes. Another aspect of dynamics of membrane proteins is their conformational flexibility. Lipids may induce the optimal conformation for catalytic activity. Breaks in Arrhenius plots of membrane-bound enzymes may be related to lipid fluidity: the break could occur when a limiting viscosity is reached for catalytic activity. Viscosity can affect protein conformational changes by inhibiting thermal fluctuations to the inner core of the protein molecule.
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PMID:Lipid fluidity and membrane protein dynamics. 332 33

Hydroxymethylglutaryl-CoA reductase (HMGR) regulates the synthesis of mevalonic acid (MVA), the precursor of the myriad of isoprenoid compounds functional in plant cells, with phytosterols representing one class of major importance. Recently, it has shown possible to solubilize and purify the membrane-bound enzyme from a heavy membrane fraction (P 16,000 x g) isolated from a cell-free homogenate of etiolated radish seedlings. What is presently known about the molecular and kinetic properties of radish HMGR is reported. Mevinolin, a highly specific competitive inhibitor of HMGR, has been valuable as a research tool in studying the regulatory role of HMGR activity for the growth and development of intact seedlings and cell cultures. The results obtained indicate a primary effect of mevinolin on phytosterol accumulation, whereas other endproducts of the multibranched isoprenoid pathway, such as ubiquinone in the mitochondria or chlorophylls and carotenoids in the plastids, are less or not at all affected. This and other data can be interpreted to mean that the organelles are autonomous in their capacity to synthesize MVA. Since the mevinolin-induced drop in free sterol accumulation is paralleled by significant plant growth retardation, a rate-limiting role of HMGR activity for phytosterol synthesis and normal development of plants is suggested.
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PMID:Hydroxymethylglutaryl-CoA reductase, a key enzyme in phytosterol synthesis? 395 69

Thenoyltrifluoroacetone (TTA) and carboxin inhibit soluble ubiquinone-deficient succinate: ubiquinone reductase according to the mixed type (with respect to added Q2) inhibition. pattern. The Ki values for the inhibitors are mutually dependent, thus indicating the presence of a single binding site for both TTA and carboxin. The enolic form of TTA was shown to be the species interacting with the enzyme. Carboxin prevents the alkali-induced inactivation of the membrane-bound succinate dehydrogenase without having any effect on the reconstitution of succinate: ubiquinone reductase from the soluble dehydrogenase and b-c1 complex. The reduction of the respiratory chain by succinate protects succinate dehydrogenase against inactivation (solubilization) by alkali; under these conditions, carboxin does not affect the inactivation process. The cumulative data suggest that the degree of the mutual mobility of the succinate dehydrogenase smaller subunit and ubiquinone reactivity-conferring protein (QPs) is a prerequisite for the catalytic mechanism of succinate: ubiquinone reductase. A mechanism of the enzyme inhibition by TTA and carboxin is proposed, which consists in non-covalent cross-linking of the subunits by the inhibitors.
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PMID:[Interaction of mitochondrial succinate:ubiquinone reductase with thenoyltrifluoroacetone and carboxin]. 399 1

Saccharomyces cerevisiae was grown in batch culture over a wide range of oxygen concentrations, varying from the anaerobic condition to a maximal dissolved oxygen concentration of 3.5 muM. The development of cells was assayed by measuring amounts of the aerobic cytochromes aa(3), b, c, and c(1), the cellular content of unsaturated fatty acids and ergosterol, and the activity of respiratory enzyme complexes. The half-maximal levels of membrane-bound cytochromes aa(3), b, and c(1), were reached in cells grown in O(2) concentrations around 0.1 muM; this was similar to the oxygen concentration required for half-maximal levels of unsaturated fatty acid and sterol. However, the synthesis of ubiquinone and cytochrome c and the increase in fumarase activity were essentially linear functions of the dissolved oxygen concentration up to 3.5 muM oxygen. The synthesis of the succinate dehydrogenase, succinate cytochrome c reductase, and cytochrome c oxidase complexes showed different responses to changes in O(2) concentration in the growth medium. Cyanide-insensitive respiration and P(450) cytochrome content were maximal at 0.25 muM oxygen and declined in both more anaerobic and aerobic conditions. Cytochrome c peroxidase and catalase activities in cell-free homogenates were high in all but the most strictly anaerobic cells.
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PMID:Respiratory development in Saccharomyces cerevisiae grown at controlled oxygen tension. 435 79

Three ubiquinone-deficient mutants of Escherichia coli unable to convert 4-hydroxybenzoate into 3-octaprenyl-4-hydroxybenzoate were isolated and examined. The results of genetic analysis suggest that each of the mutants carries a mutation in a gene designated ubiA which can be represented at minute 79 on the E. coli chromosome map. The conversion of 4-hydroxybenzoate into 3-octaprenyl-4-hydroxybenzoate, catalyzed by 4-hydroxybenzoate octaprenyltransferase, was studied with a strain of E. coli that is blocked in the common pathway of aromatic biosynthesis and consequently accumulates the precursor of the side chain of ubiquinone. Both the side-chain precursor and 4-hydroxybenzoate octaprenyltransferase were shown to be membrane-bound. The enzyme required Mg(2+) for optimal activity. The ubiA(-) mutants were found to lack 4-hydroxybenozate octaprenyltransferase activity, which suggested that the ubiA gene is the structural gene coding for this enzyme.
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PMID:Biochemical and genetic studies on ubiquinone biosynthesis in Escherichia coli K-12:4-hydroxybenzoate octaprenyltransferase. 455 89

The rates of the oxidized (Eox) and reduced (Ered) (by NAD . H through the ubiquinone pool) succinate dehydrogenase inhibition by N-ethyl-maleimide are equal and obey pseudo-first order kinetics. The protection of the enzyme against irreversible alkylation was used to quantitate the dissociation constants for Eox and Ered complexes with fumarate, succinate and malonate under conditions when no intramolecular redox reactions might occur. the membrane-bound succinate dehydrogenase catalyzes the succinate : phenazine-methosulphate reductase reaction in the presence of thenoyltrifluoroacetone by a Slater-Bonner mechanism. A comparison of the constants measured by the protection with those derived from the steady-state kinetics shows that succinate affinity for Eox is about 10 times higher than that for Ered; the reverse relations were found for fumarate, whereas the affinity for malonate only slightly depends on the redox state of the enzyme. The data obtained suggest that the dicarboxylate binding at the active site induces changes in the enzyme redox potential. The surface charge does not contribute significantly to the energy of the dicarboxylate binding to the active site of the membrane-bound enzyme.
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PMID:[Dissociation constants of succinate dehydrogenase complexes with succinate, fumarate and malonate]. 672 18

The equilibrium and rate constants for interaction of the reduced and oxidized membrane-bound succinate dehydrogenase (EC 1.3.99.1) with oxaloacetate were determined. The 10-fold decrease in the oxaloacetate affinity for the reduced enzyme was shown to be due to the 10-fold increase of the enzyme-inhibitor complex dissociation rate, which occurs upon its reduction. The rate of dissociation induced by succinate is 10 times higher than that induced by malonate in the submitochondrial particles, being equal in the soluble enzyme preparations. The rates of dissociation induced by malonate excess, or by the enzyme irreversibly utilizing oxaloacetate (transaminase in the presence of glutamate) are also equal. The data obtained suggest that succinate dehydrogenase interaction with succinate and oxaloacetate results from the competition for a single dicarboxylate-specific site. In submitochondrial particles all succinate dehydrogenase molecules are in redox equilibrium provided for by endogenous ubiquinone. No electronic equilibrium between the individual enzyme molecules exists, when succinate dehydrogenase is solubilized.
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PMID:[Interaction of succinate dehydrogenase and oxaloacetate]. 673 63


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