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: KEGG:D02011 (FAD)
5,530 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An NADH dehydrogenase possessing a specific activity 3-5 times that of membrane-bound enzyme was obtained by extraction of Acholeplasma laidlawii membranes with 9.0% ethanol at 43 degrees C. This dehydrogenase contained only trace amounts of iron (suggesting an uncoupled respiration), a flavin ratio of 1:2 FAD to FMN and 30-40% lipid. Its resistance to sedimentation is probably due to the high flotation density of the lipids. It efficiently utilized ferricyanide, menadione and dichlorophenol indophenol as electron acceptors, but not O2, ubiquinone Q10 or cytochrome c. Lineweaver-Burk plots of the dehydrogenase were altered to linear functions upon extraction with 9.0% ethanol. A secondary site of ferricyanide reduction could not be explained by the presence of cytochromes, which these membranes lack. In comparison to other respiratory chain-linked NADH dehydrogenases in cytochrome-containing respiratory chains, this dehydrogenase was characterized by similar Km's with ferricyanide, dichlorophenol indophenol, menadione as electron acceptors, but considerably smaller V's with ferricyanide, dichlorophenol indophenol, menadione as electron acceptors, and smaller specific activities. It was not stimulated or reactivated by the addition of FAD, FMN, Mg2+, cysteine or membrane lipids, and was less sensitive to respiratory inhibitors than unextracted enzyme. The ineffectiveness of ADP stimulation on O2 uptake, the insensitivity to oligomycin and the very low iron content of A. laidlawii membranes were considered in relation to conservation of energy by these cells. Some kinetic properties of the dehydrogenation, the uniquely high glycolipid content and apparently uncoupled respiration at Site I were noteworthy characteristics of this NADH dehydrogenase from the truncated respiratory chain of A. laidlawii.
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PMID:The reduced nicotinamide adenine dinucleotide "oxidase" of Acholeplasma laidlawii membranes. 17 76

The soluble hydrogenase (hydrogen: NAD+ oxidoreductase, EC 1.12.1.2) from Alcaligenes eutrophus H 16 was purified 68-fold with a yield of 20% and a final specific activity (NAD reduction) of about 54 mumol H2 oxidized/min per mg protein. The enzyme was shown to be homogenous by polyacrylamide gel electrophoresis. Its molecular weight and isoelectric point were determined to be 205 000 and 4.85 respectively. The oxidized hydrogenase, as purified under aerobic conditions, was of high stability but not reactive. Reductive activation of the enzyme by H2, in the presence of catalytic amounts of NADH, or by reducing agents caused the hydrogenase to become unstable. The purified enzyme, in its active state, was able to reduce NAD, FMN, FAD, menaquinone, ubiquinone, cytochrome c, methylene blue, methyl viologen, benzyl viologen, phenazine methosulfate, janus green, 2,6-dichlorophenoloindophenol, ferricyanide and even oxygen. In addition to hydrogenase activitiy, the enzyme exhibited also diaphorase and NAD(P)H oxidase activity. The reversibility of hydrogenase function (i.e. H2 evolution from NADH, methyl viologen and benzyl viologen) was demonstrated. With respect to H2 as substrate, hydrogenase showed negative cooperativity; the Hill coefficient was n = 0.4. The apparent Km value for H2 was found to be 0.037 mM. The absorption spectrum of hydrogenase was typical for non-heme iron proteins, showing maxima (shoulders) at 380 and 420 nm. A flavin component could be extracted from native hydrogenase characterized by its absorption bands at 375 and 447 nm and a strong fluorescense at 526 nm.
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PMID:Purification and properties of soluble hydrogenase from Alcaligenes eutrophus H 16. 18 26

Heart muscle mitochondria with satisfactory functional parameters of oxidative phosphorylation and with morphologically intact structure were isolated from canine myocardium employing a modified KEA-medium (0.18 M KCl, 10 mM EDTA, 0.5% bovine serum albumin, pH 7.1) according to Sordahl and Schwartz (1). The functional behaviour of mitochondria was investigated after different durations of in situ ischemia (cardioplegia, 15 degrees C) and correlated with metabolic findings. During ischemia the following changes were seen: 1. Successive reduction of electron flow. 2. Relatively small impairment of phosphorylation efficiency. 3. Less damage of FAD- than NAD-catalyzed oxidative phosphorylation. 4. A marked increase of electron flow and thus recovery of phosphorylation rate even after longer ischemic periods by addition of cytochrome c. As important factors of accelerating mitochondrial impairment during ischemia the myocardial ATP decrease, the lactate and H+-activity increase are discussed.
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PMID:Functional behaviour of isolated heart muscle mitochondria after in situ ischemia. Polarographic analysis of mitochondrial oxidative phosphorylation. 20 84

NADH-cytochrome c reductase, a component of benzoate 1,2-dioxygenase system, was purified to homogeneity, as judged by sodium dodecyl sulfate disc gel electrophoresis and ultracentrifugation, from benzoate-induced cells of Pseudomonas arvilla. The molecular weight of the enzyme was determined to be 38,300 by sedimentation equilibrium analysis, 37,000 by Sephadex G-100 gel filtration, and 37,500 by sodium dodecyl sulfate disc gel electrophoresis, respectively, indicating that the enzyme consisted of a single polypeptide chain. The sedimentation coefficient was calculated to be 3.3 S. The Stokes radius for the enzyme was calculated to be 27 A. The isoelectric point of the enzyme was estimated to be pH 4.2. The enzyme contained 1 mol of FAD, 2 mol of iron, and 2 mol of labile sulfide/mol of enzyme. It exhibited absorption spectrum with maxima at 273, 340, 402, and 467 nm. Amino acid analysis of the enzyme revealed that it was devoid of tryptophan. The enzyme contained 9 mol of cysteine/mol of enzyme but no disulfide linkage. The turnover number of the enzyme for the NADH-dependent reduction of cytochrome c was 17,100 at 24 degrees C. Although NADPH also acted as an electron donor, NADH was highly superior to NADPH. Ferricyanide and 2,6-dichlorophenolindophenol served as electron acceptors. Certain other properties of the enzyme are also presented.
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PMID:Characterization of NADH-cytochrome c reductase, a component of benzoate 1,2-dioxygenase system from Pseudomonas arvilla c-1. 21 33

A procedure is described for the purification of a soluble flavohemoprotein from the hydrogen bacterium Alcaligenes eutrophus. The isolated protein exists as a monomer with a molecular weight of approx. 43,000. The molecule contains two prosthetic groups, 1 mol each of noncovalently bound FAD and protoheme per monomer. The absorption spectra of the protein in its ferric, ferrous-deoxy and ferrous-carboxy forms are similar to those of hemoglobins, with the exception of the flavin contribution (absorption maxima--ferric form: 395, 456, 483, 645 nm; ferrous-deoxy form: 436, 560 nm; ferrour-CO form: 423, 539, 569 nm). The flavohemoprotein when reduced by NADH in aerobic solution is capable of binding oxygen reversibly. The stable oxygenated complex exhibits absorption maxima at 414, 541, and 576 nm. The protein catalyzes the reduction of various dyes and cytochrome c by NADH.
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PMID:An oxygen-binding flavohemoprotein from Alcaligenes eutrophus. 21 34

1. A new two-step purification is described that routinely yields 100mg quantities of component C for biochemical studies. 2. Chemical analyses show component C purified by this procedure to contain 2 g-atoms of iron, 2 mol of acid-labile sulphide (S) and 1 mol of FAD per mol of protein. 3. The Fe-S core of component C was extruded by treating the protein with p-methoxybenzenethiol in hexamethyl phosphoramide/50mM-Tris/HCl buffer, pH 8.5 (4:1, v/v), under anaerobic conditions. The spectral properties of the extruded core suggest that component C contains 1 mol of [2Fe-2S(S-Cys)4] centre per mol of protein. 4. E.p.r. spectroscopy confirms the presence of a Fe-S centre in component C. 5. Component C catalyses the reduction by NADH of ferricyanide, 2,6-dichlorophenol-indophenol or horse heart cytochrome c, with specific activities of 50--230 units/mg of protein. 6. The optimum pH for the NADH-acceptor reductase activity is 8.5--9.0, and the apparent Km values for NADH and NADPH are 0.05mM and 15.5mM respectively. 7. Unlike methane mono-oxygenase activity, NADH-acceptor reductase activity of component C is not inhibited by 8-hydroxyquinoline or by acetylene.
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PMID:Characterization of the second prosthetic group of the flavoenzyme NADH-acceptor reductase (component C) of the methane mono-oxygenase from Methylococcus capsulatus (Bath). 22 Sep 53

Adenylyl sulfate reductase has been purified from the anaerobic sulfate-reducing bacterium, Desulfovibrio vulgaris, and judged to be homogenous by several criteria. Different forms of the enzyme could be visualized in polyacrylamide gels after electrophoresis and these polymeric species have been studied by a combination of absorption spectra, polyacrylamide gel electrophoresis, and sedimentation velocity experiments. A dimeric species of molecular weight 440,000 is stable in potassium phosphate buffer but can be dissociated to a 220,000 molecular weight species by either changing the buffer system to Tris-maleate or addition of AMP, DAMP, or adenylyl sulfate. Other catalytically active nucleotides are not capable of effecting this dissociation. The enzyme was determined to contain 12 non-heme irons, 12 acid-labile sulfides, and 1 FAD per molecule when calculated on the basis of a monomeric molecular weight of 220,000. ;el electrophoresis in the presence of sodium dodecyl sulfate indicated subunits of molecular weight 72,000 and 20,000. The extinction coefficient when determined in phosphate buffer at 372 nm is 108,000 M-1 cm-a. Steady state kinetic experiments employing ferricyanide, cytochrome c, and reduced methyl viologen as artificial electron transfer reagents were performed and the kinetic constants obtained under various conditions. Several nucleotide substrates were employed and compared in each assay with respect to Km and Vmax. The reduction of cytochrome c was found to be sensitive to both anaerobiosis and superoxide dismutase, suggesting the involvement of superoxide anions with this electron acceptor.
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PMID:Some physical and kinetic properties of adenylyl sulfate reductase from Desulfovibrio vulgaris. 23 33

NADPH-cytochrome P-450 reductase was isolated from liver microsomes of phenobarbital-induced rats. The enzyme exhibits an apparent minimal molecular weight of 76,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and contains 1 molecule each of FMN and FAD. Trypsin treatment of the reductase yields an enzyme with an apparent minimal molecular weight of 69,000 which retains the ability to reduce cytochrome c but has no activity toward cytochrome P-450. Various spectrophotometric titrations were performed to examine the electron-accepting properties of the purified NADPH-cytochrome P-450 reductase and, in particular, to determine the oxidation state of the stable semiquinone form produced by air oxidation of NADPH-reduced enzyme. Titration of the air-stable semiquinone form of the reductase with ferricyanide indicated that 1 mol/2 mol of flavin was required for complete oxidation. Furthermore, a spectrum corresponding to that of the air-stable semiquinone form was produced by the addition of approximately 0.5 mol of reductant/2 mol of flavin when the oxidized enzyme was titrate with NADPH or dithionite under anaerobic conditions. The spectral changes which accompanied the overall reduction of oxidized enzyme to the reduced form with dithionite produced four sets of isosbestic points, and the spectrophotometric titration curve consisted of four approximately equal phases. In the titration with NADPH, no significant further reduction was observed after the addition of approximately 1.5 mol/2 mol of flavin. However, the enzyme was fully reduced by NADPH when an NAPH-generating system was used to prevent the accumulation of NADP. Our results establish that the air-stable semiquinone form is a 1-electron-reduced form, rather than a half-reduced (2-electron-reduced) form as maintained by others and are in agreement with earlier studies (Iyanagi, T., Makino, N., and Mason, H.S. (1974) Biochemistry 13, 1701-1710) with the purified trypsin-solubilized reductase. Accordingly, the air-stable species represents a form of the NADPH-cytochrome P-450 reductase in which one of the two flavins exists in the semiquinone state and the other in the oxidized state.
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PMID:Purified liver microsomal NADPH-cytochrome P-450 reductase. Spectral characterization of oxidation-reduction states. 63 95

Quinones constituting the electron transfer systems in a marine unicellular diatom, Phaeodactylum tricornutum, were isolated and identified chromatographically. The alga contained five quinones, i.e., plastoquinone A, plastoquinone C, plastoquinone D, alpha-tocopherylquinone, and ubiquinone-9. Other types of quinones, such as vitamin K1, were not detected. The contents of plastoquinone A, plastoquinone C, plastoquinone D, ubiquinone-9, and alpha-tocopherylquinone were 25.5, 4.95, 1.99, 4.78, and 0.28 mmol per mol of chlorophyll, respectively. The contents of the soluble C-type cytochromes, cytochrome c-550 and cytochrome c-553, were 2.15 and 4.34 mmol (heme basis) per mol of chlorophyll, respectively. The amount of B-type cytochrome in the bound form was estimated to be 3.24 mmol (heme basis) per mol of chlorophyll. The acid-soluble flavins, FAD and FMN, were present in amounts of 0.68 and 0.41 mmol per mol of chlorophyll, respectively.
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PMID:Studies on electron transfer systems in the marine diatom Phaeodactylum tricornutum. II. Identification and determination of quinones, cytochromes, and flavins. 67 Jan 58

A membrane-bound NADH dehydrogenase, solubilized and partially purified from a marine bacterium Photobacterium phosphoreum, contains FAD as the prosthetic group, and is specific for NADH. Ferricyanide, various other redox dyes and cytochrome c can act as electron acceptors. The enzymatic activity when assayed with electron acceptors other than cytochrome c, is activated by monovalent cations (Na+ and K+) and deactivated by high concentrations of monovalent anions (SCN-, NO3-, and Cl-) but not by phosphate ions. The enzymatic reaction follows a ping-pong mechanism and kinetic analysis of the enzyme showed that the activation by monovalent cations is due to increase of affinity of the enzyme for substrates; Vm was not affected. The increase of affinity was 62- and 46-fold for NADH and 57- and 31-fold for 2,6-dichlorophenol indophenol in the presence of Na+ and K+, respectively. On the other hand, NADH-cytochrome c reductase activity of the enzyme was strongly inhibited by these cations.
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PMID:Properties and kinetics of salt activation of a membrane-bound NADH dehydrogenase from a marine bacterium Photobacterium phosphoreum. 72 93


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