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: EC:1.6.99.3 (diaphorase)
5,903 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Nitric oxide (NO) may play a central role in controlling renal hemodynamics and renal salt excretion. Thus, several investigations focused on localization and function of nitric oxide synthase (NOS) isoforms in the mammalian kidney. Although studies of amphibians have contributed significantly to the elucidation of renal physiology, NOS has not been investigated in the amphibian kidney. Therefore, we characterized NOS and reduced nicotinamide adenine dinucleotide phosphate (NADPH) diaphorase biochemically and, furthermore, visualized putative NO-producing cells in the kidney of the clawed frog, Xenopus laevis. Our results indicate that NADPH-diaphorase activity correlates with NOS activity. Both enzyme activities eluted at 225 mM NaCl on a diethylaminoethanol anion exchange column and had an apparent molecular weight of 235 kDa, as estimated on an S-300 Sephacryl column. In addition, these enzymes were sensitive to Ca2+ and NADPH, but insensitive to calmodulin antagonists (trifluoperazine, W-13) or omission of calmodulin from the reaction medium. The molecular identity of NOS in Xenopus kidney extract was estimated using polymerase chain reaction. Primers to Xenopus neuronal NOS hybridized to a transcript in Xenopus kidney homogenate. NADPH-diaphorase histochemistry revealed staining in the neck segment, distal tubules, collecting segment, and peritoneal funnels. NOS-immunoreactive material was visualized in distal tubules. These results indicate that Xenopus kidney contains at least neuronal NOS, but may contain an additional NOS isoform, which is less calmodulin sensitive.
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PMID:NADPH-diaphorase activity and nitric oxide synthase activity in the kidney of the clawed frog, Xenopus laevis. 1099 86

The action of nitric oxide (NO) and the distribution of putative nitric oxide synthase-containing cells in the pelagic pteropod mollusc Clione limacina were studied using nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) histochemistry and conventional microelectrode techniques in the isolated central nervous system and in semi-intact preparations. The majority of NADPH-d-reactive neuronal somata were restricted to the cerebral ganglia. The labeled cells were small in diameter (20-30 microm) and were located in the medial areas of the ganglia. A pair of symmetrical neurons was found in the peripheral "olfactory organ." NADPH-d-reactive non-neuronal cells were detected in the periphery and were mainly associated with secretorylike cells and organs of the renopericardial system. The NO donor, diethylamine NO complex sodium salt (10-100 microM), activated neurons from both feeding and locomotory circuits. The cGMP analog, 8-Br-cGMP, mimicked the effects of NO on neurons. We suggest that NO is an endogenous neuromodulator involved in the control of some aspects of feeding and locomotor behavior of Clione.
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PMID:Distribution of NADPH-diaphorase reactivity and effects of nitric oxide on feeding and locomotory circuitry in the pteropod mollusc, Clione limacina. 1105 93

Rhodococcus sp. RB1 was able to thrive in media with up to 0.9 M NaCl or KCl and in the presence of high concentrations of nitrate (up to 0.9 M) and nitrite (up to 60 mM), but only under oxic conditions. An adaptation period was not required for salt tolerance, but a rapid extrusion of K+ and intake of Na+ was observed after addition of 0.5 M NaCl. Nitrate assimilation was limited by the carbon supply, but nitrite was not accumulated in the culture medium, even at nitrate concentrations as high as 0.8 M, thus suggesting that nitrite reduction does not limit nitrate assimilation. The presence of NaCl or KCl did not affect nitrate or nitrite uptake, which were completely inhibited by ammonium or glutamine. Rhodococcus sp. RB1 nitrate reductase had an apparent molecular mass of 142 kDa and used NADH and reduced bromophenol blue or viologens as electron donors, independently of the presence of salt. The enzyme was associated with an NADH-diaphorase activity and was induced by nitrate and repressed by ammonium or glutamine, thus showing typical biochemical and regulatory properties of bacterial assimilatory NADH-nitrate reductases. The enzyme was active in vitro in the presence of 3 M NaCl or KCI, but the maximal activity was observed at 0.5 M salt. Addition of 2 M NaCl increased the optimal temperature of the enzyme from 12 to 32 degrees C, but the optimal pH (10.3) was unaffected.
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PMID:Rhodococcus sp. RB1 grows in the presence of high nitrate and nitrite concentrations and assimilates nitrate in moderately saline environments. 1149 Oct 84

Mineralocorticoids (MC) play an important role in development of salt appetite. Part of this effect involves the hypothalamic paraventricular (PVN) and supraoptic (SON) nuclei, in which MC treatment increases arginine vasopressin (AVP) synthesis and release. Since the AVP system is also modulated by nitric oxide (NO), we studied if deoxycorticosterone acetate (DOCA) treatment changed the number of nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) active neurons and neuronal NO synthase (nNOS)-immunoreactive (IR) cells in the PVN and SON. After four injections of DOCA (10 mg/rat per day), rats developed a salt appetite and increased NADPH-d active and nNOS-IR neurons in both nuclei. A single DOCA injection did not change salt consumption or nNOS-IR cells, but increased the number of NADPH-d positive neurons in the PVN only. Therefore, while acute MC treatment stimulated the activity of pre-existing enzyme, chronic steroid treatment recruited additional neurons showing nNOS immunoreactivity/NADPH-d activity. These data suggest a role for NO produced in the PVN and SON in DOCA stimulatory effects on AVP mRNA and salt appetite.
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PMID:Deoxycorticosterone stimulates the activity of nicotinamide adenine dinucleotide phosphate-diaphorase/nitric oxide synthase immunoreactivity in hypothalamic nuclei of rats. 1218 45

Complex I is one of the respiratory chain enzymes related to NADH dehydrogenase and is an encoded gene product derived from both nuclear and mitochondrial genomes. Transcription levels of ND1 (mitochondrial) and 51 kDa (nuclear) subunits of complex I in the postnatal development of the intrinsic muscle in rat tongues were determined by Northern blot analysis. Enzyme activity levels were determined by NADH staining with tetrazolum salt, and oxygen consumption of NADH-O2 oxidoreductase activity using a Clark-type electrode. The detailed structure of the mitochondria was observed using electron microscopy. The cross-sectional area of the mitochondria gradually increased during postnatal development, and the cristae also became complex, despite the length of mitochondria in muscle fibre being constant. The mitochondria density increased from birth to 15 days of age, and declined slightly afterwards. This pattern of density resembled that of NADH-O2 oxidoreductase activity. The level of mRNA for ND1 through Northern blot analysis gradually increased from birth to 15 days of age and was highest at 21 days. For 51 kDa, the level was highest at 0 days and fell thereafter to a constant low. This suggests that the production of NADH dehydrogenase is limited by 51 kDa of Complex I derived from nuclear genomes rather than by the increase in mitochondria and composition of muscle fibre types due to changes in feeding behaviour.
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PMID:NADH-O2 oxidoreductase activity and mRNA expression of complex I (51 kDa, ND1) in postnatal intrinsic muscle of rat tongue. 1264 70

A histochemical method is described for the localization of triphosphopyridine nucleotide diaphorase using a recently synthesized tetrazolium salt (Nitro-BT). By virtue of the favorable histochemical properties of this reagent, it has been possible to demonstrate that whereas DPN diaphorase is usually restricted to the mitochondria, the TPN diaphorase activity of corresponding cells was distributed throughout the cytoplasm in granules too fine to be considered mitochondria. Furthermore, although the diaphorase alone is responsible for the passage of electrons from TPNH to the tetrazole, it has been found that sites of activity of different TPN-linked dehydrogenases can be visualized in tissue sections, and characteristic loci for each enzyme may be observed. For example, whereas TPN diaphorase and isocitric dehydrogenase have an extensive distribution in the kidney cortex, 6-phosphogluconic dehydrogenase is limited to the cells of the macula densa.
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PMID:The histochemical localization of triphosphopyridine nucleotide diaphorase. 1356 53

Cytochemical methods involving metal chelation of the formazan of an N-thiazol-2-yl tetrazolium salt are described for the localization of diphosphopyridine nucleotide diaphorase (DPND) and triphosphopyridine nucleotide diaphorase (TPND) in mitochondria. These methods utilize the reduced coenzymes DPNH or TPNH as substrate. The reaction involves a direct transfer of electrons from reduced coenzyme to the respective diaphorase which in turn transfers the electrons to tetrazolium salt, reducing it to the insoluble formazan. Competition for electrons by preferential acceptors in the respiratory chain was prevented by various inhibitors. In the presence of respiratory inhibitors the rate of tetrazolium reduction was markedly increased. The greatest reduction was observed when amytal was used. Sites of diaphorase activity appeared as deposits of blue-black metal formazan chelate measuring 0.2 to 0.3 micro in diameter. Small mitochondria contained 2 deposits, while larger ones contained up to 6. Considerable differences were observed in the rate of tetrazolium reduction and cellular localization of diaphorase activity when DPNH was used as substrate as compared to TPNH. In each instance DPNH was oxidized more rapidly by tissues than TPNH. These findings support the concept that the oxidation of coenzymes I and II is mediated through separate diaphorases.
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PMID:The cytochemical localization of oxidative enzymes. I. Diphosphopyridine nucleotide diaphorase and triphosphopyridine nucleotide diaphorase. 1361 Sep 39

Methods are presented for the intramitochondrial localization of various diphosphopyridine nucleotide and triphosphopyridine nucleotide-linked dehydrogenases in tissue sections. The cytochemical reactions studied involve the oxidation of the substrates by a specific pyridino-protein. The electron transfer of tetrazolium salt is mediated by the diaphorase system associated with the dehydrogenase. The final electron acceptor was either p-nitrophenyl substituted ditetrazole (nitro-BT) or N-thiazol-2-yl monotetrazole (MTT), the latter giving rise to metal formazan in the presence of cobaltous ions. Mitochondrial localization of the formazan precipitate could be achieved by using hypertonic incubating media containing high concentrations of substrate and co-enzyme. A fast reduction of tetrazolium salt was obtained by chemically blocking the respiratory chain enzymes beyond the flavoproteins. Although diaphorase systems are implicated in the reduction of tetrazolium salts, specific dehydrogenases are solely responsible for the distinct distribution pattern obtained in tissues with various substrates. The present findings in tissue sections are discussed in conjunction with existing biochemical evidence from differential centrifugation experiments.
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PMID:The cytochemical localization of oxidative enzymes. II. Pyridine nucleotide-linked dehydrogenases. 1361 Sep 40

A histochemical method for demonstration of glyceraldehyde-3-phosphate dehydrogenation by tissues is described. The method utilizes Nitro BT as an indicator, glyceraldehyde-3-phosphate obtained from hydrolysis of commercially obtainable glyceraldehyde-3-phosphate diethylacetal (monobarium salt) as substrate, and (ethylenediamine)tetraacetic acid acid disodium as an activating agent in a medium buffered to pH 7.2 by 0.2 M sodium phosphate. The heat lability, substrate and coenzyme specificity, and sulfhydryl and phosphate dependence of the tissue component catalyzing this reaction indicate that glyceraldehyde-3-phosphate dehydrogenase activity is being demonstrated. The disparity between the known pH optimum of this enzyme and that determined histochemically, and the anomalous histochemical localization to mitochondria of this enzyme which has been found in the soluble fraction by differential centrifugation, are thought to result from the diaphorase dependence of the tetrazolium methods and to emphasize the need for caution in the interpretation of histochemically determined intracellular localization of dehydrogenating enzymes. The evidence gathered by previous workers concerning the feasibility of demonstrating specific dehydrogenases with Nitro BT, and the correspondence of the distribution of glyceraldehyde-3-phosphate dehydrogenase determined histochemically with available quantitative data, suggest that at the cellular level the histochemical results accurately reflect the distribution of this enzyme.
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PMID:The histochemical demonstration of glyceraldehyde-3-phosphate dehydrogenase activity. 1371 13

The effects of altered dietary salt intake and/or hydralazine-induced hypotension on renal endothelial nitric oxide synthase (eNOS) expression were determined in angiotensin type-1a receptor gene knockout (At1a-/-) and wild-type (At1a+/+) mice. In At1a-/- mice, the levels of renal cortical eNOS mRNA and protein were 5 times and 3.5 times higher, respectively, in the high-salt (4% NaCl) group than in the low-salt group (0.3% NaCl). Systemic BP of the high-salt group (105 +/- 4.4 mmHg) was significantly higher than that of the low-salt group (77.0 +/- 4.7 mmHg). When hydralazine was administered to the mutant mice fed a high-salt diet, BP was reduced to 72.5 +/- 1.3 mmHg, with decreases in the levels of renal eNOS mRNA and protein expression to about half of those found in nontreated group. Consistent with the results for eNOS mRNA and protein expression, nicotinamide adenine dinucleotide phosphate (NADPH) diaphorase activity and eNOS immunoreactivity localized in the endothelium of the renal vasculature changed parallel with the amount of salt intake. In contrast to mutant mice, At1a+/+ mice did not show any changes in renal eNOS expression during the manipulation of salt intake and/or hydralazine-induced hypotension. These results suggest that At1a receptor-mediated inputs play critical roles in maintaining renal vascular eNOS expression and activity during changes in salt-water balance and systemic BP.
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PMID:Alterations in renal endothelial nitric oxide synthase expression by salt diet in angiotensin type-1a receptor gene knockout mice. 1521 63


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