Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.6.99.3 (diaphorase)
5,903 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The water-soluble carbodiimide, N-ethyl-3-(3-dimethylaminopropyl)carbodiimide was found to effectively cross-link ferredoxin to ferredoxin-NADP+ reductase. The covalent complex has a stoichiometry of 1 mol of ferredoxin per mol of the reductase. The flavoprotein moiety of the cross-linked complex maintains most of its diaphorase activity and more interestingly has gained the capacity to catalyze the NADPH-cytochrome c reaction without addition of free ferredoxin in the assay mixture. Furthermore, the cross-linked complex binds NADP+ with a Kd = 88 microM at an ionic strength of 0.02 M. These results show that a ternary complex among the reductase and its substrates can be formed, suggesting that the binding sites for ferredoxin and the pyridine nucleotides are distinct. The bound ferredoxin can interact with cytochrome c; the iron-sulfur cluster of the cross-linked complex is shown to be reduced under anaerobic conditions by NADPH and to be required for the catalysis of the NADPH-cytochrome c reductase reaction. The cross-linked complex, added to thylakoids inhibited by the antibody against the reductase, catalyzes the H2O-cytochrome c photoreduction, which suggests that the ferredoxin moiety of the complex can interact with its electron donor in the photosynthetic chain. Restoration of NADP+ photoreduction requires the addition of free ferredoxin.
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PMID:A cross-linked complex between ferredoxin and ferredoxin-NADP+ reductase. 672 48

The non-ionic detergent lauryl dimethylamine N-oxide (LDAO) has been used to extract the NADH dehydrogenases of Arum maculatum mitochondria. Affinity chromatography on 5'-ADP-Sepharose 4B was used to separate the rotenone-sensitive (complex I) NADH dehydrogenase from the rotenone-insensitive NADH dehydrogenase. An 18-fold purification of the rotenone-insensitive NADH dehydrogenase was achieved. The enzyme is specific for NADH with optimal activity around pH 7.2. The apparent Km for NADH is 28 microM, with dichloroindophenol as acceptor at pH 7.2. The rotenone-insensitive NADH dehydrogenase appears to be a flavoprotein and no iron-sulphur centres were detected by electron spin resonance spectroscopy.
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PMID:Purification and characterization of the rotenone-insensitive NADH dehydrogenase of mitochondria from Arum maculatum. 674 60

The diurnal rhythms of the microsomal flavoprotein NADPH-cytochrome c reductase activity, of diaphorase and of succinic dehydrogenase are presented. Minimum levels are ascertained at 09(00), maximum levels at 21(00). The concentration of mitochondrial radicals as a function of the time of day is also demonstrated. Here too the minimum is at 09(00) and the maximum between 15(00) and 21(00). On the other hand, GSH levels are found to be high between 09(00) and 12(00) and low in the evening. Thus a causative relationship between the concentration of cellular radicals, which originate in flavin enzymes, and the concentration of the tripeptide glutathione is assumed.
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PMID:Flavin enzymes, mitochondrial radicals and reduced glutathione in daily rhythmic dependency. 677 1

The triazine dyes, Cibacron blue F3GA and Procion red HE3B inhibited diaphorase activity of ferredoxin-NADP+ reductase, in a competitive manner with respect to NADPH. The Ki values were 1.5 and 0.2 microM, respectively. Binding of the dyes to the flavoprotein, as measured by difference spectroscopy, indicated an apparent stoichiometry of 1 mol dye/mol reductase and was prevented by NADP+ or high ionic strength. Chemical modification of a lysine residue and a carboxyl group at the NADP(H) binding site of the enzyme prevented complex formation with Procion red. Procion red showed a higher affinity for ferredoxin-NADP+ reductase than Cibacron blue. The Kd values were 1.9 and 5 microM, respectively. Once covalently linked to a Sepharose matrix, the triazine compounds specifically bind the flavoprotein. The interaction is partially electrostatic and partially hydrophobic. The enzyme can be eluted by high concentrations of salt or low concentrations of the corresponding coenzyme. The use of this affinity column allows the rapid purification of ferredoxin-NADP+ oxidoreductase from spinach leaves with good yields.
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PMID:Interaction of ferredoxin-NADP+ oxidoreductase with triazine dyes. A rapid purification method by affinity chromatography. 682 90

The purification and properties of metlegoglobin reductase from lupine (Lupinus luteus L.) nodules are described. The purification procedure results in a 1056-fold purification of the enzyme with a total yield of 21%. The enzyme possesses the NADH-diaphorase activity. Metlegoglobin reductase is heterogenous during electrophoresis and isoelectric focusing. Electrophoresis produces two vicinal active bands, while isoelectrofocusing results in four active fractions. The fraction possessing the highest activity has a pI of 4.4. The enzyme is a flavoprotein, in which all flavins are represented by FAD. The molecular weight of the enzyme is 30 000. In some properties metlegoglobin reductase from lupine nodules is similar to methemoglobin reductase from erythrocytes and metmyoglobin reductase from muscles.
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PMID:[Properties of metlegoglobin reductase from lupine nodules]. 689 54

1. A reflection spectrometric method was developed which allowed the simultaneous measurement of flavoprotein absorption and fluorescence on an in vitro preparation of brown adipose tissue. 2. From their spectral characteristics and from the effects of substrates and a metabolic inhibitor (amytal) it was shown that the absorption and fluorescence signals are associated with different flavoproteins. 3. The fluorescence signal is mainly due to changes in the redox state of NADH dehydrogenase, and the absorption signal to changes in redox state of he flavoproteins in the acyl-CoA dehydrogenase pathway. 4. The results suggest that changes in the flavoprotein redox state in response to electrical nerve stimulation, exogenous norepinephrine and substrate addition reflect changes in the metabolic activity of the tissue. These responses were studied in the postnatal period. 5. The amplitude of the tissue response to either nerve stimulation or norepinephrine administration is already maximal at birth and decreases in animals 50 days old. The frequency of nerve stimulation of the concentration of norepinephrine required to produce a half maximum response is significantly higher for the new-born as compared to 13 day and 50 day old animals. 6. For small stimulation intensities a steady state oxidation of the NADH dehydrogenase concomitant with a steady state reduction of the flavoproteins in the acyl-CoA dehydrogenase pathway was recorded. 7. It is concluded that in rats less than 12 hours old, brown adipose tissue is functionally innervated although previous histochemical studies had failed to detect nerve terminals containing catecholamines at this early age.
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PMID:Postnatal development of sympathetic innervation of rat brown adipose tissue reevaluated with a method allowing for monitoring flavoprotein redox state. 713 28

Mitochondrial NADH dehydrogenase may be isolated from bovine heart as a lipoprotein complex (Complex I or NADH-ubiquinone oxidoreductase). Polypeptide subunits that are exposed to the hydrophobic region of the phospholipid bilayer were identified by photolabelling with the hydrophobic probe, 5-[125I]iodonaphth-1-yl azide. Chaotropic resolution of the labelled enzyme showed that the hydrophilic flavoprotein and iron-protein fragments of the enzyme were not in contact with the phospholipid bilayer. When complex I that had been partially depleted of phospholipids was photolabelled, incorporation of radioactivity into certain polypeptides was increased, indicating either conformational changes in protein or preferential association of these polypeptides with residual cardiolipin. A model NADH dehydrogenase structure is proposed on the basis of these results and those obtained with hydrophilic probes by Smith & Ragan (1980) Biochem. J. 185, 315-326.
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PMID:Identification of the subunits of bovine heart mitochondrial NADH dehydrogenase that are exposed to the phospholipid bilayer by photo-labelling with 5-iodonaphth-1-yl azide. 723 4

The purification by affinity chromatography up to homogeneity and the properties of NAD-reductase from purple sulfur bacterium Thiocapsa roseopersicina, strain BBS, are described. The molecular weight of NAD-reductase is about 80000; pI is 3.9. The enzyme consists of two subunits. According to the stabilizing effect of FAD at preparative electrophoresis and the inhibitory effect of atebrine NAD-reductase is a flavoprotein. The bulk of the enzyme (about 75%) is localized in the cell periplasmic space. NAD-reductase is less thermostable and has a lower O2 stability as compared to the NADP-reductase from the same organism. The enzyme is specific to NADH ane catalyzes the menadione-reductase reaction, diaphorase reaction of benzyl viologen and methyl viologen reductions. In the presence of NADH NAD-reductase reduces cytochromes c552 and "c3" from T. roseopersicina and forms a complex with spinach ferredoxin.
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PMID:[Purification and properties of NAD-reductase from phototrophic bacterium Thiocapsa roseopersicina]. 723 99

NADH-cytochrome c reductase, a component of benzoate 1,2-dioxygenase system, is an ion-sulfur flavoprotein with one FAD and one iron-sulfur cluster of [2Fe-2S] type (Yamaguchi, M., and Fujisawa, H. (1978) J. Biol. Chem. 253, 8848-8853). Treatment of NADH-cytochrome c reductase with p-chloromercuriphenylsulfonic acid resulted in fading of its color with a concomitant loss of the NADH-cytochrome c reductase activity. The p-chloromercuriphenylsulfonic acid-treated enzyme was found to contain one FAD but no significant amounts of iron and labile sulfide. Incubation of the iron-sulfur-depleted enzyme with ferrous ions and sulfide in the presence of 2-mercaptoethanol led to both reconstitution of iron-sulfur cluster of the enzyme and concomitant restoration of the enzyme activity. Although the iron-sulfur-depleted enzyme catalyzed NADH-dependent reduction of ferricyanide, nitroblue tetrazolium, or oxygen, it could not catalyze NADH-dependent reduction of the oxygenase component of benzoate 1,2-dioxygenase system. In contrast, the reconstituted enzyme recovered the activity of NADH-dependent reduction of the oxygenase component to the original level. Certain other catalytic and molecular properties of the iron-sulfur-depleted enzyme and the reconstituted enzyme are also presented.
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PMID:Reconstitution of iron-sulfur cluster of NADH-cytochrome c reductase, a component of benzoate 1,2-dioxygenase system from Pseudomonas arvilla C-1. 724 Feb 44

The role of flavins in vitamin K function was assessed by examining blood coagulation and in vitro activities of hepatic vitamin K-dependent enzymes from control and riboflavin-deficient rats. One-stage prothrombin times and Factor VII activities were lower in flavin-deficient rats than in ad libitum or pair-fed controls. Fibrinogen, prothrombin, and Factor X activities were normal. Hepatic vitamin K-dependent carboxylase activity was severely depressed in flavin-deficient rats when assayed with [vitamin K + NADH] and somewhat depressed with reduced vitamin K (vitamin KH2) as substrate. One-hour flavin repletion appreciably restored [vitamin K + NADH]-dependent activity, but vitamin KH2-dependent activity was not restored even after 16 hours repletion. These results suggest that the carboxylating enzyme itself is not a flavoprotein, but that the microsomal NADH dehydrogenase required for [vitamin K + NADH]-dependent carboxylation is a flavoprotein. This dehydrogenase may differ from the cytosolic Warfarin-inhibitable 'DT-diaphorase' in that the activity of the latter, which is reduced 50% in flavin-deficient rats, is not at all restored by one-hour flavin repletion. Flavin status-dependent differences in NADH-dependent or vitamin KH2-dependent epoxidation of vitamin K paralleled differences in the carboxylase. Flavin deficiency had no effect on vitamin K 2,3-epoxide reductase activity nor on its inhibition by Warfarin.
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PMID:Vitamin K-dependent reactions in rat liver: role of flavoproteins. 731 May 34


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