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)

Disruption of the PDX1 gene encoding the protein X component of the mitochondrial pyruvate dehydrogenase (PDH) complex in Saccharomyces cerevisiae did not affect viability of the cells. However, extracts of mitochondria from the mutant, in contrast to extracts of wild-type mitochondria, did not catalyze a CoA- and NAD(+)-linked oxidation of pyruvate. The PDH complex isolated from the mutant cells contained pyruvate dehydrogenase (E1 alpha + E1 beta) and dihydrolipoamide acetyltransferase (E2) but lacked protein X and dihydrolipoamide dehydrogenase (E3). Mutant cells transformed with the gene for protein X on a unit-copy plasmid produced a PDH complex that contained protein X and E3, as well as E1 alpha, E1 beta, and E2, and exhibited overall activity similar to that of the wild-type PDH complex. These observations indicate that protein X is not involved in assembly of the E2 core nor is it an integral part of the E2 core. Rather, protein X apparently plays a structural role in the PDH complex; i.e., it binds and positions E3 to the E2 core, and this specific binding is essential for a functional PDH complex. Additional evidence for this conclusion was obtained with deletion mutations. Deletion of most of the lipoyl domain (residues 6-80) of protein X had little effect on the overall activity of the PDH complex. This observation indicates that the lipoyl domain, and its covalently bound lipoyl moiety, is not essential for protein X function. However, deletion of the putative subunit binding domain (residues approximately 144-180) of protein X resulted in loss of high-affinity binding of E3 and concomitant loss of overall activity of the PDH complex.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Disruption and mutagenesis of the Saccharomyces cerevisiae PDX1 gene encoding the protein X component of the pyruvate dehydrogenase complex. 200 23

NAD(P)H quinone oxidoreductase (DT diaphorase; EC 1.6.99.2) activity in cultured Syrian hamster fibroblastic cells was measured. The cells examined were classified into three categories: (1) four primary embryonic fibroblasts, (2) three non-malignant immortal cells and (3) four malignantly transformed cells. The results showed that cytosolic DT diaphorase activity in malignant cells was markedly higher than the corresponding activity in non-malignant cells (enzyme activity: (3) much greater than (1) greater than or equal to (2)). The enzyme activity was not influenced very much by the particular phase of cell growth. Thus, the increase in DT diaphorase activity seems to be a good marker for malignant transformation, at least in Syrian hamster fibroblastic cells.
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PMID:Marked increases in DT diaphorase activity in malignantly transformed Syrian hamster fibroblastic cells. 202 24

The authors studied the cytotoxic function, activity of NAD- and NADP-diaphorases in the peripheral blood lymphocytes in 57 patients with B-cellular variant of chronic lympholeukoses and found a significant reduction of the natural killer activity of lymphocytes, increased activity of NADP-diaphorase. Reduction of natural killer activity in patients with B-cell variant of chronic lympholeukoses did not depend on the activity of membrane diaphorases in peripheral blood lymphocytes.
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PMID:[NAD- and NADP-diaphorase activity in the peripheral blood lymphocytes of patients with the B-cell variant of chronic lympholeukemia]. 204 48

The NAD(P)H- and flavoprotein fluorescence signals of the isolated perfused rat kidney were characterized by their electrochemical properties and by comparing them with those of isolated kidney cortex mitochondria. From titration experiments with lactate pyruvate under anoxic conditions it is concluded that about 25% of the NAD(P)H-fluorescence can be attributed to the cytosolic pyridine nucleotides. For this signal a midpoint potential of -268 mV was calculated. The major part of the NAD(P)H-fluorescence signal was of mitochondrial origin, showing a midpoint potential of -304 mV which was titrated with beta-hydroxybutyrate/acetoacetate. A comparable value (Em7.4 = -310 mV) was found for isolated kidney cortex mitochondria using the same redox couple. The flavoprotein fluorescence could be totally attributed to the mitochondrial compartment and was found to originate from the NAD-dependent alpha-lipoamide dehydrogenase and the electron transfer flavoprotein of beta-oxidation to approx. 60% and 40%, respectively. A midpoint potential of -285 mV was determined for the NAD-dependent part of the flavoprotein signal. On the basis of these results it can be concluded that the fluorescence changes which were caused by ouabain in the normoxically perfused rat kidney reflect an increase in reduction of the mitochondrial NAD-system.
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PMID:[Characterization of surface fluorescence signals of isolated perfused rat kidney]. 209 17

By crossed immunoelectrophoresis with antibodies against the NAD-linked hydrogenase the presence of three hydrogenase protein species was demonstrated in crude extracts of Alcaligenes eutrophus H16. Protein 1 (antigen 1) exhibited NAD-reducing activity and was shown to be identical with the native heterotetrameric enzyme. Protein 2 (antigen 2) was catalytically inactive in the antibody-precipitated form and corresponded to the beta subunit (56 kDa) of the holoenzyme. Protein 3 (antigen 3) was serologically distinct from antigen 2 and catalyzed NADH-oxidizing (diaphorase) activity, suggesting that it either consists of the alpha peptide or of the alpha and gamma subunits of the diaphorase dimer. Tandem immunoelectrophoresis revealed that antigen 2 was the predominant protein species in cells cultivated under nickel deficiency. Low concentrations of the diaphorase-active antigen 3 were also detected under these conditions. Extracts from mutants defective in the catalytic activity of NAD-reducing hydrogenase still contained the four polypeptides. This was shown by immunodiffusion and immunoblotting with antibodies raised against the individual subunits. However, as observed with nickel-deficient cells, no complete tetrameric protein could be identified, and the dominant subunit species (70-80%) was the beta peptide.
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PMID:Identification of distinct NAD-linked hydrogenase protein species in mutants and nickel-deficient wild-type cells of Alcaligenes eutrophus H16. 211 62

This study reports a new property of the important NAD(P)H-dependent flavoenzymes, glutathione reductase, lipoyl dehydrogenase and ferredoxin-NADP+ oxidoreductase, that can catalyze a one electron reduction of metal ions such as chromium(VI) and vanadium(V). During the enzymatic reduction process, molecular oxygen is reduced to H2O2, which reacts with the reduced metal complexes to generate hydroxyl radicals. Since the hydroxyl radicals have been suggested to play an important role in Cr(VI) toxicity, this study provides a basis for a recent observation that Cr(VI) mutagenesis is strongly oxygen dependent. These results also point to an enzymatic pathway for the metabolism of some metal ions and concomitant generation of hydroxyl radicals.
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PMID:NADPH-dependent flavoenzymes catalyze one electron reduction of metal ions and molecular oxygen and generate hydroxyl radicals. 217 63

The DNA sequence of the Salmonella typhimurium ahp locus was determined. The locus was found to contain two genes that encode the two proteins (C22 and F52a) that comprise the S. typhimurium alkyl hydroperoxide reductase activity. The predicted sequence of the F52a protein component of the alkyl hydroperoxide reductase was found to be highly homologous to the Escherichia coli thioredoxin reductase protein (34% identity with many conservative substitutions). The homology was found to be particularly striking in the region containing the redox-active cysteines of the thioredoxin reductase molecule, and among the identities were the redox-active cysteines themselves. Aside from the strong similarity to thioredoxin reductase, overall homology between the F52a protein and other flavoprotein disulfide oxidoreductases such as glutathione reductase, dihydrolipoamide dehydrogenase, and mercuric reductase was found to be rather limited, and the conserved active site segment common to the three proteins was not observed within the F52a protein. However, three short segments that have been implicated in FAD and NAD binding were found to be conserved between the F52a protein and the other disulfide reductases. These results suggest that the alkyl hydroperoxide reductase is the second known member of a class of disulfide oxidoreductases which was represented previously by thioredoxin reductase alone; they also allow the putative assignment of several functional domains.
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PMID:Alkyl hydroperoxide reductase from Salmonella typhimurium. Sequence and homology to thioredoxin reductase and other flavoprotein disulfide oxidoreductases. 219 51

The activity of human myocardial enzymes in sudden coronary death (SCD) was quantitatively histochemically examined. The activity of succinate dehydrogenase (SDH), lactate dehydrogenase (LDH), beta-oxybutyrate dehydrogenase (beta-OBDH), alpha-glycerolphosphate dehydrogenase (alpha-GPDH), NAD-diaphorase (NAD-ase), and glucose-6-phosphate dehydrogenase (G-6-PDH) was measured on prompt autopsies (up to 3 hours of death onset). beta-OBDH and LDH showed an increase in activity in the myocardium from the subjects who had suddenly died from coronary heart disease without evident changes in the heart. In SCD in the presence of small cardiosclerosis, the activity of the enzymes characterizing the major processes of energy generation was also enhanced, which was caused by moderately severe myocardial hypertrophy. In the myocardium from the subjects who had died from coronary heart disease in the presence of large postinfarction cardiosclerosis, the activity of the enzymes was directly related to the degree of myocardial hypertrophy and the signs of chronic heart failure. As myocardial hypertrophy progressed, the enzymatic activity rose, but there were signs of chronic heart failure, it fell. The findings suggest that the changes in myocardial enzymatic activity in SCD are heterogeneous and associated with the type of prior abnormalities in the cardiovascular system.
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PMID:[Disorders of myocardial metabolism in sudden coronary death in the presence of coronary atherosclerosis: findings of quantitative histoenzymologic studies]. 221 37

Three different dihydrolipoamide dehydrogenases were purified to homogenity from the anaerobic glycine-utilizing bacteria Clostridium cylindrosporum, Clostridium sporogenes, and Peptostreptococcus glycinophilus, and their basic properties were determined. The enzyme isolated from P. glycinophilus showed the properties typical of dihydrolipoamide dehydrogenases: it was a dimer with a subunit molecular mass of 53,000 and contained 1 mol of flavin adenine dinucleotide and 2 redox-active sulfhydryl groups per subunit. Only NADH was active as a coenzyme for reduction of lipoamide. Spectra of the oxidized enzyme exhibited maxima at 230, 270, 353, and 453 nm, with shoulders at 370, 425, and 485 nm. The dihydrolipoamide dehydrogenases of C. cylindrosporum and C. sporogenes were very similar in their structural properties to the enzyme of P. glycinophilus except for their coenzyme specificity. The enzyme of C. cylindrosporum used NAD(H) as well as NADP(H), whereas the enzyme of C. sporogenes reacted only with NADP(H), and no reaction could be detected with NAD(H). Antibodies raised against the dihydrolipoamide dehydrogenase of C. cylindrosporum reacted with extracts of Clostridium acidiurici, Clostridium purinolyticum, and Eubacterium angustum, whereas antibodies raised against the enzymes of P. glycinophilus and C. sporogenes showed no cross-reaction with extracts from 42 organisms tested.
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PMID:Purification and comparative studies of dihydrolipoamide dehydrogenases from the anaerobic, glycine-utilizing bacteria Peptostreptococcus glycinophilus, Clostridium cylindrosporum, and Clostridium sporogenes. 229 86

An enzymatic assay method for the determination of urinary formic acid is described. Formic acid in urine was cleaved to carbon dioxide and water by formic acid dehydrogenase, whereby NAD+ was converted to NADH, which reacted with INT (p-iodonitrotetrazolium violet) in the presence of NAD-diaphorase. The color thus produced was determined at 500 nm. In addition, a simple gas chromatographic method of urinary formic acid is described, in which head space gas of formic acid methylester was applied into the wide bore column. The urinary formic acid concentrations by the enzymatic method agreed well with that by the gas chromatographic method. A simple gas chromatographic method for urinary methanol assay is also described. Acetonitrile was added to an equal volume of urine containing methanol. After centrifugation, the supernatant was injected into gas chromatography (GC). The peaks of urinary methanol and ethanol were separated by GC. Formic acid and methanol in urine of unexposed healthy subjects and workers exposed to methanol were analyzed by the colorimetric and gas chromatographic methods. Geometric mean concentrations of urinary formic acid and methanol in the healthy subjects were 7.82 mg/g creatinine and 1.34 mg/l, respectively. The concentration ratio of formic acid to methanol in the urine of the workers exposed to methanol was calculated to be 3.67 +/- 2.10, which agreed with the ratio under a controlled exposure experiment. A slower excretion of formic acid than that of methanol in the urine of a volunteer was also observed.
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PMID:Enzymatic assay of formic acid and gas chromatography of methanol for urinary biological monitoring of exposure to methanol. 234 46


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