Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.6.99.1 (NADPH-diaphorase)
 3,903 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The objective of this study was to determine whether nitric oxide (NO) is produced locally in the bovine corpus luteum (CL) and whether NO mediates prostaglandin F2alpha (PGF2alpha)-induced regression of the bovine CL in vivo. The local production of NO was determined in early I, early II, mid, late, and regressed stages of CL by determining NADPH-d activity and the presence of inducible and endothelial NO synthase immunolabeling. To determine whether inhibition of NO production counteracts the PGF2alpha-induced regression of the CL, saline (10 ml/h; n = 10) or a nonselective NOS inhibitor (Nomega-nitro-l-arginine methyl ester dihydrochloride [L-NAME]; 400 mg/h; n = 9) was infused for 2 h on Day 15 of the estrous cycle into the aorta abdominalis of Holstein/Polish Black and White heifers. After 30 min of infusion, saline or cloprostenol, an analogue of PGF2alpha (aPGF2alpha; 100 microg) was injected into the aorta abdominalis of animals infused with saline or L-NAME. NADPH-diaphorase activity was present in bovine CL, with the highest activity at mid and late luteal stages (P < 0.05). Inducible and endothelial NO synthases were observed with the strongest immunolabeling in the late CL (P < 0.05). Injection of aPGF2alpha increased nitrite/nitrate concentrations (P < 0.01) and inhibited P4 secretion (P < 0.05) in heifers that were infused with saline. Infusion of L-NAME stimulated P4 secretion (P < 0.05) and concomitantly inhibited plasma concentrations of nitrite/nitrate (P < 0.05). Concentrations of P4 in heifers infused with L-NAME and injected with aPGF2alpha were higher (P < 0.05) than in animals injected only with aPGF2alpha. The PGF2alpha analogue shortened the cycle length compared with that of saline (17.5 +/- 0.22 days vs. 21.5 +/- 0.65 days P < 0.05). L-NAME blocked the luteolytic action of the aPGF2alpha (22.6 +/- 1.07 days vs. 17.5 +/- 0.22 days, P < 0.05). These results suggest that NO is produced in the bovine CL. NO inhibits luteal steroidogenesis and it may be one of the components of an autocrine/paracrine luteolytic cascade induced by PGF2alpha.
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PMID:Administration of a nitric oxide synthase inhibitor counteracts prostaglandin F2-induced luteolysis in cattle. 1260 17

1. Enzyme systems from Cucurbita pepo have been shown to catalyse the reduction of nitrite and hydroxylamine to ammonia in yields about 90-100%. 2. Reduced benzyl viologen serves as an efficient electron donor for both systems. Activity of the nitrite-reductase system is directly related to degree of dye reduction when expressed in terms of the function for oxidation-reduction potentials, but appears to decrease to negligible activity below about 9% dye reduction. 3. NADH and NADPH alone produce negligible nitrite loss, but NADPH can be linked to an endogenous diaphorase system to reduce nitrite to ammonia in the presence of catalytic amounts of benzyl viologen. 4. The NADH- or NADPH-nitrate-reductase system that is also present can accept electrons from reduced benzyl viologen, but shows relationships opposite to that for the nitrite-reductase system with regard to effect of degree of dye reduction on activity. The product of nitrate reduction may be nitrite alone, or nitrite and ammonia, or ammonia alone, according only to the degree of dye reduction. 5. The relative activities of nitrite-reductase and hydroxylamine-reductase systems show different relationships with degree of dye reduction and may become reversed in magnitude when effects of degree of dye reduction are tested over a suitable range. 6. Nitrite severely inhibits the rate of reduction of hydroxylamine without affecting the yield of ammonia as a percentage of total substrate loss, but hydroxylamine has a negligible effect on the activity of the nitrite-reductase system. 7. The apparent K(m) for nitrite (1 mum) is substantially less than that for hydroxylamine, for which variable values between 0.05 and 0.9mm (mean 0.51 mm) have been observed. 8. The apparent K(m) values for reduced benzyl viologen differ for the nitrite-reductase and hydroxylamine-reductase systems: 60 and 7.5 mum respectively. 9. It is concluded that free hydroxylamine may not be an intermediate in the reduction of nitrite to ammonia by plants, and a possible mechanism for reduction of both compounds by the same enzyme system is discussed in the light of current ideas relating to other organisms.
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PMID:THE REDUCTION OF NITRATE, NITRITE AND HYDROXYLAMINE TO AMMONIA BY ENZYMES FROM CUCURBITA PEPO L. IN THE PRESENCE OF REDUCED BENZYL VIOLOGEN AS ELECTRON DONOR. 1434 47

The cnidarian nervous system is considered by many to represent neuronal organization in its earliest and simplest form. Here we demonstrate, for the first time in the Cnidaria, the neuronal localization of nitric oxide synthase (NOS) in the hydromedusa Aglantha digitale (Trachylina). Expression of specific, fixative-resistant NADPH-diaphorase (NADPH-d) activity, characteristic of NOS, was observed in neurites running in the outer nerve ring at the base of the animal and in putative sensory cells in the ectoderm covering its tentacles. At both sites, diphenyleneiodonium (10(-4) M) abolished staining. Capillary electrophoresis confirmed that the NO breakdown products NO2- and NO3- were present at high levels in the tentacles, but were not detectable in NADPH-d-negative areas. The NADPH-d-reactive neurons in the tentacles send processes to regions adjacent to the inner nerve ring where swimming pacemaker cells are located. Free-moving animals and semi-intact preparations were used to test whether NO is involved in regulating the swimming program. NO (30-50 nM) and its precursor L-arginine (1 mM) stimulated swimming, and the effect was mimicked by 8-Br-cGMP (50-100 microM). The NO scavenger PTIO (10-100 microM) and a competitive inhibitor of NOS, L-nitroarginine methyl ester (L-NAME, 200 microM), significantly decreased the swimming frequency in free-moving animals, while its less-active stereoisomer D-nitroarginine methyl ester (D-NAME, 200 microM) had no such effect. 1H-[1,2,4]oxadiazolo[4,3-a]quinoxaline-1-one (ODQ, 5-20 microM), a selective inhibitor of soluble guanylyl cyclase, suppressed spontaneous swimming and prevented NO-induced activation of the swimming program. We suggest that an NO/cGMP signaling pathway modulates the rhythmic swimming associated with feeding in Aglantha, possibly by means of putative nitrergic sensory neurons in its tentacles.
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PMID:Nitric oxide regulates swimming in the jellyfish Aglantha digitale. 1498 73

Several independent studies of bacterial degradation of nitrate ester explosives have demonstrated the involvement of flavin-dependent oxidoreductases related to the old yellow enzyme (OYE) of yeast. Some of these enzymes also transform the nitroaromatic explosive 2,4,6-trinitrotoluene (TNT). In this work, catalytic capabilities of five members of the OYE family were compared, with a view to correlating structure and function. The activity profiles of the five enzymes differed substantially; no one compound proved to be a good substrate for all five enzymes. TNT is reduced, albeit slowly, by all five enzymes. The nature of the transformation products differed, with three of the five enzymes yielding products indicative of reduction of the aromatic ring. Our findings suggest two distinct pathways of TNT transformation, with the initial reduction of TNT being the key point of difference between the enzymes. Characterization of an active site mutant of one of the enzymes suggests a structural basis for this difference.
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PMID:Biotransformation of explosives by the old yellow enzyme family of flavoproteins. 1518 58

Severely Ca-deficient Triticum aestivum L. seedlings accumulated high levels of nitrite and moderate levels of nitrate and organic nitrogen, but contained unaltered levels of hydroxylamine. Nitrite accumulation was not related to molybdenum deficiency, or altered cellular pH. Nitrate reductase was decreased by Ca deficiency, apparently by repression of enzyme synthesis from accumulated nitrite and not by inhibition of enzyme activity. Nitrite reductase and NADP diaphorase activities were not affected by Ca deficiency, and Ca did not restore activity to nitrite reductase inactivated by cyanide. The results indicated that the role of Ca is in intracellular transport of nitrite and not in induction or activity of enzymes.
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PMID:Evidence for a role of calcium in nitrate assimilation in wheat seedlings. 1665 39

Molybdenum is absolutely required for the nitrate-reducing activity of the nicotinamide adenine dinucleotide nitrate reductase complex isolated from Chlorella fusca. The whole enzyme nicotinamide adenine dinucleotide nitrate reductase is formed by cells grown in the absence of added molybdate, but only its first activity (nicotinamide adenine dinucleotide diaphorase) is functional. The second activity of the complex, which subsequently participates also in the enzymatic transfer of electrons from nicotinamide adenine dinucleotide to nitrate (FNH(2)-nitrate reductase), depends on the presence of molybdenum. Neither molybdate nor nitrate is required for nitrate reductase synthesis de novo, but ammonia acts as a nutritional repressor of the complete enzyme complex. Under conditions which exclude de novo synthesis of nitrate reductase, the addition of molybdate to molybdenum-deficient cells clearly increases the activity level of this enzyme, thus suggesting in vivo incorporation of the trace metal into the pre-existing inactive apoenzyme.Competition studies with tungstate corroborate these conclusions and indicate that the only role played by molybdenum in Chlorella is connected with the reduction of nitrate to nitrite. Tungsten seems to act by replacing molybdenum in the nitrate reductase complex, thus rendering inactive the FNH(2)-nitrate reductase portion of the nicotinamide adenine dinucleotide nitrate reductase complex.
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PMID:Role of molybdenum in nitrate reduction by chlorella. 1665 84

THE ASSIMILATORY NITRATE REDUCTASE (NADH: nitrate oxidoreductase, E.C. 1.6.6.2.) from the marine diatom Thalassiosira pseudonana, Hasle and Heimdal, has been purified 200-fold and characterized. The regulation of nitrate reductase in response to various conditions of nitrogen nutrition has been investigated.Nitrate reductase activity is repressed by the presence of ammonium in vivo, and its synthesis is derepressed when ammonium is absent. The derepression process is sensitive to cycloheximide and apparently requires protein synthesis. Repression of enzyme activity by ammonium is neither inhibited nor delayed by the presence of cycloheximide. In vitro, ammonium does not inhibit enzyme activity.NADH is the physiological electron donor for the enzyme in a flavin-dependent reaction. Spectral studies have indicated the presence of a b-type cytochrome associated with the enzyme. It is possible to observe enzymatic oxidation-reduction reactions which represent partial functions of the over-all electron transport capacity of this enzyme. Nitrate reductase will accept electrons from artificial electron donors such as reduced methyl viologen in a flavin-independent reaction. Further, dithionitereduced flavin adenine dinucleotide can donate electrons to the enzyme to reduce nitrate to nitrite. Finally, the nitrate reductase will exhibit a diaphorase activity and reduce the artificial electron acceptor mammalian cytochrome c in flavin-adeninedinucleotide-dependent reaction.Inhibition studies with potassium cyanide, sodium azide, and o-phenanthroline have yielded indirect evidence for metal component (s) of the enzyme.The inhibition of the NADH-requiring enzyme activities by p-hydroxymercuribenzoate has shown that an essential sulfhydryl group is involved in the initial portion of the electron transport. Heat treatment exerts an effect similar to the p-hydroxymercuribenzoate inhibition; namely, the NADH-requiring activities are rapidly inactivated, whereas the terminal nitrate-reducing activities are relatively stable to heat.The T. pseudonana nitrate reductase molecule has the hydrodynamic properties of an ellipsoid with a frictional coefficient of 1.69 and a molecular weight of 330,000.
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PMID:Purification and Characterization of the Nitrate Reductase from the Diatom Thalassiosira pseudonana. 1665 41

The nitrite-reducing activity of the normal susceptible biotype of lambsquarters (Chenopodium album L.) was strongly inhibited by atrazine in the assay medium, both in the case of the in vivo assays of leaf discs in light, and in vitro photoreduction assays of crude extracts. In vitro assays of crude extracts with methylviologen or ferredoxin supplying the reducing potential were not inhibited by atrazine. In the resistant biotype, inhibition of nitrite reduction did not occur with any of the above assays. Thus, it appears that atrazine does not inhibit nitrite reductase itself, but rather the availability of photosynthetically supplied electrons for the reduction. Atrazine had no effect when added to the media for either in vivo or in vitro assays of nitrate reduction by either the susceptible or resistant biotype.Young lambsquarters plants were treated with atrazine by spraying the leaves at a rate which was lethal for susceptible plants after 5 or 6 days, but had little effect on the resistant biotype. Nitrite did not accumulate in either biotype, but remained present at the level of about 0.1 microgram nitrite N per gram fresh weight. The nitrate content of susceptible-type leaves did increase to two or three times the initial level, during the first four days after spraying. Usually the only visible effect on the plants during this time was a decreased growth rate. Twenty-four hours after spraying the following activities had fallen to 25% or less of the activities of solvent-sprayed control plants: in vivo nitrite reductase, in vivo nitrate reductase, in vitro NADH-nitrate reductase, in vitro reduced flavin mononucleotidenitrate reductase, and in vitro NADH-diaphorase. In these atrazine-treated plants, in vitro nitrite reductase activity with reducing potential supplied by methylviologen was not affected, nor were any of the above activities in leaves of atrazine-treated resistant plants. The abrupt fall in nitrate reductase represents an effect of atrazine not directly related to inhibition of photosynthesis.
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PMID:Reduction of Nitrate and Nitrite in Lambsquarters (Chenopodium album) Biotypes Resistant and Susceptible to Atrazine Toxicity. 1666 20

All nitrate reductase-related activities of Chlamydomonas reinhardtii wild-type and mutant 305 cells were degraded in vivo under conditions in which the reversible inactivation could take place. When the enzyme was in the inactive form, half-lives of all nitrate reductase-related activities in wild and mutant 305 strains decreased significantly. The only nitrate reductase-related activity present in mutant 104, nitrate reductase-diaphorase, was incapable of undergoing reversible inactivation and was not degraded under any of the conditions tested. Addition of nitrate to inactive nitrate reductase of mutant 305 caused the in vivo reactivation of the enzyme and halted its degradation. Our results indicate that reversibly inactivated nitrate reductase from C. reinhardtii is the main target for a degradation system, and that nitrate reductase related diaphorase must be integrated in a reversibly inactive nitrate reductase complex to undergo degradation. A physiological role for the interconversion process of nitrate reductase can be understood on the basis of these facts.
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PMID:Involvement of Reversible Inactivation in the Regulation of Nitrate Reductase Enzyme Levels in Chlamydomonas reinhardtii. 1666 99

Cultures of Lemna gibba L. G3 were maintained at a constant, N-limited growth rate by adding nitrate daily in amounts calculated to sustain a rate of culture N increment of 0.20 day(-1). Nitrate added to the culture was consumed within 8 to 10 hours and the partitioning to reduction and accumulation during this phase corresponded to, on the average, 75 and 25% of net uptake, respectively. The calculated rate of nitrate reduction was stimulated by onset of net uptake without delay and decreased when net uptake ceased. NADH-nitrate reductase (NR) activity measured in vitro without inclusion of antiproteolytic agents more than doubled during the first hour after nitrate addition and then gradually fell to its original level over the rest of the 24 hour interval. In the presence of the proteinase inhibitor leupeptin during extraction, however, NR activity was in general much higher and without any apparent cycles. The relative stabilizing effect of leupeptin was greatest on NADH-NR and reduced flavin adenine mononucleotide-NR activities whereas the effect was less on NADH-cytochrome c reductase activity (diaphorase) and reduced methylviologen-NR activity. The constant nitrate reductase activity measured in the presence of proteinase inhibitors is assumed to reflect the physiological situation. It thus appeares that short-term changes in nitrate assimilation by N-limited Lemna is related to the flux of nitrate to the reducing site and not to changes in nitrate reductase activity.
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PMID:Nitrogen Utilization in Lemna: I. Relations between Net Nitrate Flux, Nitrate Reduction, and in Vitro Activity and Stability of Nitrate Reductase. 1666 90


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