Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: DrugBank:EXPT00568 (ascorbate)
23,072 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Chemical-modification studies on submitochondrial particle pyridine dinucleotide transhydrogenase (EC 1.6.1.1) demonstrate the presence of one class of sulfhydryl group in the nicotinamide adenine dinucleotide phosphate (NADP) site and another peripheral to the active site. Reaction of the peripheral sulfhydryl group with N-ethylmaleimide, or both classes with 5,5'-dithiobis(2-nitrobenzoic acid), completely inactivated transhydrogenase. NADP+ or NADPH nearly completely protected against 5,5'-dithiobis(2-nitrobenzoic acid) inactivation and modification of both classes of sulfhydryl groups, while NADP+ only partially protected against and NADPH substantially stimulated N-ethylmaleimide inactivation. Methyl methanethiolsulfonate treatment resulted in methanethiolation at both classes of sulfhydryl groups, and either NADP+ or NADPH protected only the NADP site group. S-Methanethio and S-cyano transhydrogenases were active derivatives with pH optima shifted about 1 unit lower than that of the native enzyme. These experiments indicate that neither class of sulfhydryl group is essential for transhydrogenation. Lack of involvement of either sulfhydryl group in energy coupling to transhydrogenation is suggested by the observations that S-methanethio transhydrogenase is functional in (a) energy-linked transhydrogenation promoted by phenazine methosulfate mediated ascorbate oxidation and (b) the generation of a membrane potential during the reduction of NAD+ by reduced nicotinamide adenine dinucleotide phosphate (NADPH).
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PMID:Chemical modification of mitochondrial transhydrogenase: evidence for two classes of sulfhydryl groups. 3

Membrane vesicles isolated from cells of bacillus subtilis W23 accumulate manganese in the presence of an energy source. The artificial electron donor system ascorbate and phenazine methosulfate or reduced nicotinamide adenine dinucleotide and phenazine methosulfate can supply the energy for the uptake. D-Lactate in the presence or absence of phenazine methosulfate would not support manganese accumulation. Anaerobiosis, cyanide, m-chlorophenyl carbonylcyanide hydrozone, valinomycin, gramicidin, and p-hydroxy-mercuribenzoate inhibit the uptake. The inhibition by p-hydroxymercuribenzoate is prevented by excess dithiothreitol. Potassium fluoride or sodium arsenate has no effect on the uptake. The manganese transport system in the B. subtilis vesicles exhibits Michaelis-Menten kinetics with a Km of 13 muM and a Vmax of 1.7 nmol/min per mg (dry weight) of membranes. The uptake of manganese is specific and is not inhibited by 0.1 mM CaCL2 or Mgcl2.
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PMID:Active transport of manganese in isolated membrane vesicles of Bacillus subtilis. 4 50

The degradation of DNA by bleomycin was studied in the absence and in the presence of added reducing agents, including 2-mercaptoethanol, dithiothreitol, reduced nicotinamide adenine dinucleotide phosphate, H2O2, and ascorbate, and in the presence of a superoxide anion generating system consisting of xanthine oxidase and hypoxanthine. In all cases, breakage of DNA was inhibited by low concentrations of chelators; where examined in detail, deferoxamine mesylate was considerably more potent than (ethylenedinitrilo)tetraacetic acid. Iron was found to be present in significant quantities in all reaction mixtures. Thus, the pattern of inhibition observed is attributed to the involvement of contaminating iron in the degradation of DNA by bleomycin. Cu(II), Zn(II), and Co(II) inhibit degradation of DNA by bleomycin and Fe(II) in the absence of added reducing agents. A model is proposed in which the degradation of DNA in these systems is dependent on the oxidation of an Fe(II)-bleomycin-DNA complex.
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PMID:Effect of chelating agents and metal ions on the degradation of DNA by bleomycin. 8 Feb 26

Membrane vesicles of Veillonella alcalescens, grown in the presence of L-lactate and KNO-3, actively transport amino acids under anaerobic conditions in the presence of several electron donors and the electron acceptor nitrate. The highest initial rates of uptake are obtained with L-lactate, followed by reduced nicotinamide adenine dinucleotide, glycerol-1-phosphate, formate, and L-malate.. The membrane vesicles contain the dehydrogenases for these electron donors, and these enzymes are coupled with nitrate reductase. In membrane vesicles from cells, grown in the presence of nitrate, the dehydrogenases are not coupled with fumarate reducatase, and anaerobic transport of amino acids does not occur with fumarate as electron acceptor. Under aerobic conditions none of the physiological electron donors can energize transport. However, a high rate of uptake is observed with the electron donor system ascorbate-phenazine metho-sulfate. This electron donor system also effectively energizes transport under anaerobicconditions in the presence of the electron acceptor nitrate.
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PMID:Amino acid transport in membrane vesicles of obligately anaerobic Veillonella alcalescens. 16 33

Membrane vesicles isolated from Bacillus subtilis W23 catalyze active transport of the C4 dicarboxylic acids L-malate, fumarate, and succinate under aerobic conditions in the presence of the electron donor reduced beta-nicotinamide adenine dinucleotide or the non-physiological electron donor system ascorbate-phenazine methosulfate. The dicarboxylic acids are accumulated in unmodified form. Inhibitors of the respiratory chain, sulfhydryl reagents, and uncoupling agents inhibit the accumulation of the dicarboxylic acids. The affinity constants for transport of L-malate, fumarate, and succinate are 13.5, 7.5, and 4.3 muM, respectively; these values are severalfold lower than those reported previously for whole cells. Active transport of these dicarboxylic acids occurs via one highly specific transport system as is indicated by the following observations. (i) Each dicarboxylic acid inhibits the transport of the other two dicarboxylic acids competitively. (ii) The affinity constants determined for the inhibitory action are very similar to those determined for the transport process. (iii) Each dicarboxylic acid exchanges rapidly with a previously accumulated dicarboxylic acid. (iv) Other metabolically and structurally related compounds do not inhibit transport of these dicarboxylic acids significantly, except for L-aspartate and L-glutamate. However, transport of these dicarboxylic amino acids is mediated by independent system because membrane vesicles from B. subtilis 60346, lacking functional dicarboxylic amino acid transport activity, accumulate the C4 dicarboxylic acids at even higher rates than vesicles from B. subtilis W 23. (v) A constant ratio exists between the initial rates of transport of L-malate, fumarate, and succinate in all membrane vesicle preparations isolated from cells grown on various media. This high-affinity dicarboxylic acid transport system seems to be present constitutively in B. subtilis W23.
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PMID:Dicarboxylic acid transport in membrane vesicles from Bacillus subtilis. 17 Dec 51

The membrane-bound respiratory system of the gram-negative bacterium Spirillum itersonii was investigated. It contains cytochromes b (558), c (550), and o (558) and beta-dihydro-nicotinamide adenine dinucleotide (NADH) and succinate oxidase activities under all growth conditions. It is also capable of producing D-lactate and alpha-glycerophosphate dehydrogenases when grown with lactate or glycerol as sole carbon source. Membrane-bound malate dehydrogenase was not detectable under any conditions, although there is high activity of soluble nicotinamide adenine dinucleotide: malate dehydrogenase. When grown with oxygen as the sole terminal electron acceptor, approximately 60% of the total b-type cytochrome is present as cytochrome o, whereas only 40% is present as cytochrome o in cells grown with nitrate in the presence of oxygen. Both NADH and succinate oxidase are inhibited by azide, cyanide, antimycin A, and 2-n-heptyl-4-hydroxyquinoline-N-oxidase at low concentrations. The ability of these inhibitors to completely inhibit oxidase activity at low concentrations and their effects upon the aerobic steady-state reduction levels of b- and c-type cytochromes as well as the aerobic steady-state reduction levels obtained with NADH, succinate, and ascorbate-dichlorophenolindophenol suggest that presence of an unbranched respiratory chain in S. itersonii with the order ubiquinone leads to b leads to c leads to c leads to oxygen.
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PMID:Membrane-bound respiratory of Spirillum itersonii. 18 74

The electron transport system of Neisseria gonorrhoeae was partially characterized by using spectrophotometric, spectroscopic, and oxygen consumption measurements. The effects of selected electron transport inhibitors (amytal, rotenone, 2-heptyl-4-hydroxyquinoline, antimycin A1, and potassium cyanide [KCN]) on electron transfer in whole-cell and sonically treated whole-cell preparations of N. gonorrhoeae were examined. The oxidation of reduced nicotinamide adenine dinucleotide, measured as a decrease in absorbance at 340 nm, was inhibited by each of the compounds tested. Oxygen consumption stimulated by reduced nicotinamide adenine dinucleotide was also inhibited, whereas oxygen uptake stimulated by succinate and malate was inhibited by KCN alone, suggesting the presence of a KCN-sensitive terminal oxidase. Room temperature optical difference spectra indicate an operational electron bypass around the amytal-rotenone-binding site. Difference spectra in the presence of 2-heptyl-4-hydroxyquinoline suggest a possible site of interaction of this compound at the substrate side of cytochrome b. Reduced-minus-oxidized spectra of ascorbate-tetramethyl-p-phenylenediamine suggest the participation of b-, a-, and d-type cytochromes in terminal oxidase activity. Hence, N. gonorrhoeae appears to have an electron transport chain containing cytochrome c, two b-type cytochromes (one of which has an oxidase function), and possibly a- and d-type cytochromes. An abbreviated chain exists through which succinate and malate can be oxidized directly by a KCN-sensitive component.
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PMID:Effects of selected inhibitors on electron transport in Neisseria gonorrhoeae. 20 70

Optical absorbance difference spectra of membrane vesicles prepared from aerobically grown Pseudomonas putida indicated that, when harvested in logarithmic phase, the cells contained one c-type cytochrome and two or three b-type cytochromes, one of which was cytochrome o. As the cells grew into stationary phase and the oxygen concentration of the medium dropped to essentially zero, an additional component believed to be cytochrome d was produced. Both the o- and d-type cytochromes might function as terminal oxidases. No a-type cytochromes could be detected at any stage of growth. Polarographic measurement of oxygen utilization revealed that cyanide and azide are effective inhibitors of the oxidation of ascorbate coupled with 2,6-dichlorophenolindophenol or N,N,N',N'-tetramethyl-p-phenylenediamine in respiratory particles from either log-phase or stationary-phase cells. Reduced nicotinamide adenine dinucleotide- or succinate-dependent oxygen utilization, however, was sensitive to these inhibitors only in log-phase particles. These results indicate that an alternate terminal oxidase may be synthesized by this organism in response to restricted oxygen availability and that branching of the respiratory system may result.
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PMID:Changes in cytochrome content and electron transport patterns in Pseudomonas putida as a function of growth phase. 61 38

Membrane vesicles from the menaquinone-deficient Bacillus subtilis aro D contain a low content of menaquinone and consequently oxidaze reduced nicotinamide adenine dinucleotide (NADH) at low rate. Supplementation of the membrane vesicles suspension with the menaquinone-analogue menadione, results in an incorporation of menadione in the membranes. The incorporated menadione increases with the external menadione concentration up to a maximum of 7 nmol of menadione bound per mg membrane protein. The NADH oxidase activity of the membrane vesicles increases linearly with the menadione content and a 35-fold stimulation is obtained in fully reconstituted membrane vesicles; this maximal NADH oxidase activity is about two-fold higher than the NADH oxidase activity in membrane vesicles from wild-type B.subtilis W23. Supplementation of membrane vesicles from B.subtilis W23 with menadione also results in a stimulation of the NADH oxidase activity but only a stimulation of 1.6-fold is maximally obtained. The NADH oxidase activities in reconstituted B.subtilis aro D and B.subtilis W23 membrane vesicles are similarly affected by respiratory chain inhibitors, indicating that menadione occupies physiological sites of menaquinone. NADH and the non-physiological electron donor ascorbate + phenazine methosulphate are the best energy sources for active amino acid transport in membrane vesicles from B.subtilis W23. Membrane vesicles from B.subtilis aro D accumulate amino acids in the presence of acorbate + phenazine methosulphate, but not with NADH. However, membrane vesicles from this mutant, reconstituted with menadione, demonstrate NADH-driven transport activity. This activity increases linearly with the NADH oxidase activity, but maximal transprt activities are reached under conditions where the NADH oxidase activity is not yet maximal. These results indicate that the rate of energy supply is the limiting factor for transport at low NADH oxidase activities and that the transport system itself becomes the limiting factor for transport at low NADH oxidase activities and that the transport system itself becomes the limiting factor under conditions of high NADH oxidase activities. Under energy-limiting conditions 135-235 molecules of NADH have to be oxidized in order to transport one molecule of amino acid. At all levels of energy supply a competition by the different amino acid transport systems for the available energy could not be observed. These observations indicate that only a fraction of the energy, generated by the respiratory chain, is used for the transport of an amino acid and that the bulk of the energy dissipates via other channels in the membrane vesicles.
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PMID:Reconstitution of reduced nicotinamide adenine dinucleotide oxidase activity with menadione in membrane vesicles from the menaquinone-deficient Bacillus subtilis aro D. Relation between electron transfer and active transport. 82 14

A new soluble hemoprotein, designated as H-450, has been purified from pig liver. The absolute absorption spectrum of H-450 shows maxima at 550 and 428 nm. The dithionite-reduced H-450 has absorption peaks at 572, 540, and 450 nm; the unique Soret band at 450 nm is the basis for our tentative designation of this new hemoprotein as H-450 (hemoprotein 450). The spectrum of dithionite-reduced H-450 at 77 K gives two alpha peaks (571 and 566 nm), three beta peaks (546, 537, and 529 nm), and a Soret band at 449 nm. The prosthetic group of H-450 has been identified as protoheme IX. Gel electrophoresis experiments show that H-450 is composed of two nonidentical subunits, alpha and beta (mol wts = 61 000 and 45 000). H-450 contains 1 mol of heme/alphabeta dimer of 106 000 molecular weight. Preliminary sedimentation equilibrium experiments suggest a minimum molecular weight of 218 000 for the native protein. This corresponds to a tetramer, alpha2beta2 containing two heme groups. H-450 is not reduced by reduced nicotinamide adenine dinucleotide (NADH), NADH phosphate, ascorbate, or ferrocyanide. Neither reduced nor oxidized H-450 binds CO, 1 mM cyahide, or 1 mM azide. Dithionite-reduced H-450 is autoxidizable. The molar extinction coefficient of native H-450 is 261 X 103 at 280 nm and 263 X 103 at 428 nm. The purification procedure involves homogenization, high-speed centrifugation, ammonium sulfate fractionation, diethylaminoethylcellulose chromatography, density gradient centrifugation, a calcium phosphate gel step, and a second density gradient centrifugation. The procedure yeilds approximately 2 mg of purified protein from 750 g of pig liver.
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PMID:Isolation and properties of a new, soluble, hemoprotein (H-450) from pig liver. 99 Feb 54


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