Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.8.1.4 (diaphorase)
 2,754 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A feasibility study aimed at stabilization of L-lactate-dehydrogenase, L-malate-dehydrogenase, alcohol-dehydrogenase and diaphorase by the recently described method of enzyme 'encagement' was conducted. This method involves derivatizing the enzymes with polyglutaraldehyde, followed by secondary crosslinking with amino derivatives of polyacrylamide. Encagement conditions were optimized for each of the four enzymes, so as to achieve the highest thermal stability combined with highest catalytic activity. Depending on the encagement conditions, residual activities were in the range of 18% to 96% with higher values in the presence of cofactors. Increases in thermal stabilization of up to 26-fold were obtained. For high retention of enzymic activity and stability, the most significant factor was the concentration of polyglutaraldehyde; the crosslinking polymers had only a negligible effect. Furthermore, the significant enhancement in thermal stability could be attained without perturbing the kinetic parameters: Km values for NADH and pH optima remained unaltered for the stabilized enzymes.
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PMID:Stabilization of NAD(+)-dependent dehydrogenases and diaphorase by bilayer encagement. 136 7

These studies concern the initial steps in 4-nitroquinoline 1-oxide (4NQO) metabolism in relation to mechanisms of anticarcinogenesis. Butylated hydroxyanisole (BHA) administration by a protocol known to inhibit the pulmonary tumorigenicity of 4NQO in A/HeJ mice enhanced hepatic and pulmonary activities for 4NQO metabolism by two major pathways, conjugative detoxification and nitroreductive activation. High-performance liquid chromatography analysis showed approximate doubling of two types of glutathione transferase subunits with 4NQO-conjugating activity in livers of BHA-treated mice. Similar increases were observed in hepatic 4NQO-conjugating activity and in Vmax, while Km for 4NQO was 39 to 43 microM. Pulmonary 4NQO-glutathione transferase activity increased 24 to 29%. DT diaphorase activity toward 4NQO was elevated 3.3-fold in livers and 2.7-fold in lungs of BHA-treated mice. However, the predominant 4NQO reductase of liver and lung was dicumarol resistant, had a strong preference for NADH, and showed little if any response to BHA. This Mr 200,000 enzyme, partially purified from livers of Swiss mice, exhibited the stoichiometry of 2-NADH/4NQO expected for reduction of 4NQO to 4-hydroxyaminoquinoline 1-oxide. Its high affinity for 4NQO (Km, 15 microM) signified a much greater influence on 4NQO metabolism than DT diaphorase (Km, 208 microM). The dicumarol-resistant 4NQO reductase differed from several known cytosolic nitroreductases. The results suggest that protection by BHA may result from alteration of the balance between 4NQO activation and conjugation.
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PMID:Nitroreductases and glutathione transferases that act on 4-nitroquinoline 1-oxide and their differential induction by butylated hydroxyanisole in mice. 137 76

The neuron morphology and distribution of four putative transmitters were investigated in the myenteric plexus of frog (Rana esculenta) midgut. The gross morphology was revealed by NADH-diaphorase histochemistry, and the shape of the neurons by silver impregnation. Nerve cells had heterogeneous distribution: they either formed ganglia or placed as solitary neurons in the duodenum, while in the rest of the midgut only solitary neurons were observed. Three morphologically distinct cell types were revealed by silver impregnation: mainly type I and type II neurons cells were seen in the duodenum, while the rest of the intestine contained type II and III cells. Catecholamine fluorescence was revealed in nerve fibres in the duodenum, while few small nerve cells were observed in the small intestinal region. Acetylcholinesterase histochemistry showed strongly reactive nerve cells that were associated with the main fibre bundles in the duodenum. Only longitudinally oriented fibres and occasionally stained neurons were seen in the small intestine. Substance P immunocytochemistry revealed an extensive plexus, which contained a moderate number of stained perikarya in the full length of the midgut. Gamma-aminobutyric acid showed non-uniform distribution in the two parts of the midgut: a stronger and more regular fibre staining was found in the duodenum then in the rest of the intestine. Ultrastructural observations demonstrated that intrinsic neurons received synaptic inputs from the profiles contained agranular vesicles, while "P"-type profiles established close contacts with neurons. Both profile types formed close contacts with the smooth muscle cells.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Some morphological and histochemical features of the midgut myenteric plexus of the common European frog, Rana esculenta. 137 78

Lipopolysaccharide (LPS), either alone or in combination with cytokines, induces nitric oxide (NO) synthase activity in cells that normally release little or no NO. In arterial smooth muscle cells and various macrophage cell lines, NO synthase activity is induced after several hours of incubation with LPS. In brain, NADPH-dependent diaphorase activity has been associated with constitutive NO synthase. Here we show that incubation of rat aorta or cultured macrophages with LPS causes a time-dependent induction of NO synthase. The NO synthase activity in both rat aorta and macrophages was calcium independent and inhibited by NG-monomethyl-L-arginine and NG-nitro-L-arginine. We also found that LPS caused a time-dependent induction in NADPH-dependent diaphorase activity in both rat aorta and cultured macrophages. The diaphorase activity was mainly NADPH dependent and NADH independent. NO synthase activity and NADPH-diaphorase activity in crude cytosol from LPS-treated macrophages were found to co-purify, using 2',5'-ADP-Sepharose followed by Superose-6 gel permeation chromatography.
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PMID:Induction of NADPH-dependent diaphorase and nitric oxide synthase activity in aortic smooth muscle and cultured macrophages. 137 28

The enzyme DT diaphorase (NAD(P)H dehydrogenase (quinone), EC 1.6.99.2) is unusual in that it can utilize either NADH or NADPH as a co-factor for the reduction of its substrates. We have shown that the intact NAD(P)H molecule is not required and that other reduced pyridinium compounds can also act as co-factors for DT diaphorase. The entire adenine dinucleotide portion of NAD(P)H can be dispensed with entirely and the simplest quaternary (and therefore reducible) derivative of nicotinamide, 1-methylnicotinamide, was as effective as NAD(P)H as a co-factor for the reduction of the quinone, menadione. Nicotinamide 5'-O-benzoyl riboside was also as effective a co-factor as NAD(P)H, whilst nicotinamide ribotide and riboside have a higher Km, and decreased the kcat of DT diaphorase. Nicotinic acid derivatives had little activity. Kinetic analysis indicated that both nicotinamide ribotide and riboside may be interacting with the menadione binding site rather than the NAD(P)H site. Irrespective of the differences between the various reduced pyridinium derivatives in their ability to act as co-factors for the reduction of menadione by DT diaphorase, all the compounds that showed activity in this assay were equally effective co-factors for the reduction of the nitrobenzamide, CB 1954 (5-(aziridin-1-yl)-2,4-dinitrobenzamide). The apparent Km of DT diaphorase for all these co-factors approached zero. It was concluded that co-factor binding is not a rate-limiting step in the nitroreductase activity of DT diaphorase.
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PMID:Identification of novel reduced pyridinium derivatives as synthetic co-factors for the enzyme DT diaphorase (NAD(P)H dehydrogenase (quinone), EC 1.6.99.2). 138 52

NADH was metabolized both by serum components and at the cell surface. The metabolism by serum was either oxidation to NAD+, or hydrolysis of the pyrophosphate to yield nicotinamide mononucleotide (reduced) (NMNH) and AMP. NMNH was further hydrolysed to yield nicotinamide riboside (reduced) (NRH), which was stable. NAD+ was hydrolysed (although at a slower rate than was NADH), but was also reduced to yield NADH. The reduction of NAD+ was catalysed by the enzyme serum L(+)lactate dehydrogenase (EC 1.1.1.27) and was dependent on the concentration of L(+)lactate in the serum. NADPH was hydrolysed in a similar manner to NADH but not oxidized by serum. NADH generated from NAD+ by serum derived from human, foetal calf and horse sources was capable of driving the bioreductive activation of CB 1954 by the enzyme DT diaphorase. Cell surfaces oxidized NADH to NAD+, but did not oxidize NADPH or NRH. These observations suggest that NAD(P)H would be unsuitable as a source of reducing equivalents for the bioreductive activation of prodrugs by a reductase enzyme in Antibody Directed Enzyme Prodrug Therapy (ADEPT). In contrast, NAD+ (which could act as a source of NADH) and NRH could avoid the shortcomings of NAD(P)H, and act as suitable cofactors for an enzyme in an ADEPT system.
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PMID:Metabolism of NAD(P)H by blood components. Relevance to bioreductively activated prodrugs in a targeted enzyme therapy system. 138 14

This paper describes the development of the rat vomeronasal organ from the stage of anlage until adulthood. Groups of four rats were sacrificed daily from prenatal day 13 (E13) until birth; at days 2, 4, 7, 10, 14 and 16 after birth; weekly from day P21 to P42 plus an additional group of adults. The vomeronasal organs were processed for light microscopy, including alcian blue-PAS and NADH-diaphorase reactions, and also for electron microscopy. For summarizing our results we propose the following developmental stages: 1. Anlage (E13). 2. Early morphogenesis (E14-16). 3. Late morphogenesis (E17 to birth). 4. Initiation of secretory activity (First postnatal week). 5. Cytoarchitectural maturity (2nd postnatal week). 6. Complete maturity (From 3rd postnatal week onwards). Our results on the maturation of the histological structure and the histochemical reactions, indicate that there may be some functional activity at birth but the development of the organ still continues during the first three postnatal weeks to acquire its full functional capability.
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PMID:Developmental stages of the vomeronasal organ in the rat: a light and electron microscopic study. 144 18

The toxicity of CB 1954 [5-(aziridin-1-yl)-2,4-dinitrobenzamide] towards human cells was greatly enhanced by NADH (when foetal calf serum was present in the culture medium) and by nicotinamide riboside (reduced) (NRH), but not by nicotinate riboside (reduced). Co-treatment of human cells with CB 1954 and NADH resulted in the formation of crosslinks in their DNA. The toxicity produced by other DNA crosslinking agents was unaffected by reduced nicotinamide compounds. When caffeine was included in the medium, a reduction in the cytotoxicity of CB 1954 occurred. The toxicity experienced by human cell lines after exposure to CB 1954 and NADH was proportional to their levels of the enzyme DT diaphorase NAD(P)H dehydrogenase (quinone), EC 1.6.99.2. It is concluded that NRH, which we have shown to be a co-factor for rat DT diaphorase (Friedlos et al., Biochem Pharmacol 44: 25-31, 1992), is generated from NADH by enzymes in foetal calf serum, and stimulates the activity of human DT diaphorase towards CB 1954.
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PMID:Potentiation of CB 1954 cytotoxicity by reduced pyridine nucleotides in human tumour cells by stimulation of DT diaphorase activity. 144 31

1. Kinetic studies of lipoamide dehydrogenase and its modified enzymes catalyzing lipoamide oxidoreduction and ancillary reactions at various pH are compared. 2. The asymptotic kinetics of lipoamide oxidoreductions switch between the ping pong and ordered mechanisms by varying pH of the reactions. 3. pH-rate profiles of these reactions are bell-shaped suggesting the participation of 2 ionizable residues with pK values of 6.6 +/- 0.5 and above 8 respectively. 4. The unusually high pK value for the catalytic site histidine is attributed to its involvement in an ion-pair formation. 5. In the absence of the catalytic site histidine, the pH-rate profile for the lipoamide reduction of the photooxidized enzyme is no longer bell-shaped but it is similar to those of the transhydrogenation and NADH-oxidation of the native enzyme. 6. This implies the participation of a low-pK protonated group in these reactions.
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PMID:pH dependent kinetic studies of lipoamide dehydrogenase catalysis. 145 16

In order to elucidate the mechanism of the biological activation of nitrofurans, the interaction of these compounds with lipoamide dehydrogenase (LipDH)** was investigated. LipDH catalysed one-electron reduction of several nitrofuran derivatives. The reaction could be demonstrated spectroscopically and was enhanced by cadmium, arsenite and anaerobiosis. The role of flavin in the nitroreductase activity was supported by (a) the nitrofuran effect on the spectral properties of anaerobic, arsenite-inhibited, NADH-reduced LipDH; (b) FAD catalytic activity in a NADH-nitrofuran model system; and (c) the nitroreductase activity of LipDH monomer. Two-electron nitrofuran reduction to less oxidized products was inhibited by cadmium, arsenite and NAD+. The possible role of reactive nitrosofuran derivatives as intermediates of the nitrofuran reduction sequence was supported by the LipDH capability for catalysing 2-nitroso-1-naphthol redox-cycling. The nitroso naphthol reduction was inhibited by cadmium and arsenite, like the two-electron nitrofuran reduction.
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PMID:Reduction of nitrofuran compounds by heart lipoamide dehydrogenase: role of flavin and the reactive disulfide groups. 145 54


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