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)

Liver homogenates have been submitted to quantitative fractionation by differential centrifugation. Three particulate fractions: N (nuclear), ML (large granules), and P (microsomes), and a final supernate (S) have been obtained. The biochemical composition of the microsomal fraction has been established from the assay and distribution pattern of 25 enzymatic and chemical constituents. These included marker enzymes for mitochondria (cytochrome oxidase), lysosomes (acid phosphatase and N-acetyl-beta-glucosaminidase), and peroxisomes (catalase). The microsomal preparations were characterized by a moderate contamination with large cytoplasmic granules (only 6.2% of microsomal protein) and by a high yield in microsomal components. Enzymes such as glucose 6-phosphatase, nucleoside diphosphatase, esterase, glucuronyltransferase, NADPH cytochrome c reductase, aminopyrine demethylase, and galactosyltransferase were recovered in the microsomes to the extent of 70% or more. Another typical behavior was shown by 5'-nucleotidase, alkaline phosphatase, alkaline phosphodiesterase I, and cholesterol, which exhibited a "nucleomicrosomal" distribution. Other complex distributions were obtained for several constituents recovered in significant amount in the microsomes and in the ML or in the S fraction.
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PMID:Analytical study of microsomes and isolated subcellular membranes from rat liver. II. Preparation and composition of the microsomal fraction. 415 Apr 89

Rat liver microsomal fractions have been equilibrated in various types of linear density gradients. 15 fractions were collected and assayed for 27 constituents. As a result of this analysis microsomal constituents have been classified, in the order of increasing median density, into four groups labeled a, b, c, and d. Group a includes: monoamine oxidase, galactosyltransferase, 5'-nucleotidase, alkaline phosphodiesterase I, alkaline phosphatase, and cholesterol; group b: NADH cytochrome c reductase, NADPH cytochrome c reductase, aminopyrine demethylase, cytochrome b(5), and cytochrome P 450; group c: glucose 6-phosphatase, nucleoside diphosphatase, esterase, beta-glucuronidase, and glucuronyltransferase; group d: RNA, membrane-bound ribosomes, and some enzymes probably adsorbed on ribosomes: fumarase, aldolase, and glutamine synthetase. Analysis of the microsomal fraction by differential centrifugation in density gradient has further dissociated group a into constituents which sediment more slowly (monoamine oxidase and galactosyltransferase) than those of groups b and c, and 5'-nucleotidase, alkaline phosphodiesterase I, alkaline phosphatase, and the bulk of cholesterol which sediment more rapidly (group a2). The microsomal monoamine oxidase is attributed, at least partially, to detached fragments of external mitochondrial membrane. Galactosyltransferase belongs to the Golgi complex. Group a2 constituents are related to plasma membranes. Constituents of groups b and c and RNA belong to microsomal vesicles derived from the endoplasmic reticulum. These latter exhibit a noticeable biochemical heterogeneity and represent at the most 80% of microsomal protein, the rest being accounted for by particles bearing the constituents of groups a and some contaminating mitochondria, lysosomes, and peroxisomes. Attention is called to the operational meaning of microsomal subfractions and to their cytological complexity.
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PMID:Analytical study of microsomes and isolated subcellular membranes from rat liver. 3. Subfractionation of the microsomal fraction by isopycnic and differential centrifugation in density gradients. 415 Apr 90

The biologic basis for the elevated histochemical reduction of nitroblue tetrazolium dye (NBT) in neutrophils from patients with acute bacterial infection or polycythemia vera was studied. A precipitin reaction followed mixing NBT with heparin. NBT was reduced after phagocytosis of this complex (H-NBT) by polymorphonuclear leukocytes (PMNs). Ingestion required divalent cations and was facilitated by the presence of complement. H-NBT incubated with normal but not with C2-deficient human serum converted native C3 to its inactive form. Phagocytic indices were determined in patients and controls by measuring O(2) utilization and hexose monophosphate shunt activity and by visually counting cell-associated latex particles. Significant elevations above controls were observed in phagocytes isolated from all patients with elevated histochemical NBT scores when H-NBT complex, latex, or zymosan was employed as the phagocytic particle. Increased indices were observed in the presence of fresh AB serum, heat-inactivated AB serum, or without serum. Serum from patients with elevated NBT scores did not alter phagocytosis in control phagocytes. With NADH and NADPH as substrates, total NBT diaphorase activity of sonicated leukocytes was normal in all patients. These results suggest that increased phagocytic capacity of PMNs is the primary cause of increased histochemical NBT reduction. The PMNs of patients with acute bacterial infection or polycythemia vera may have alterations in their cell membranes which lead to an enhanced rate of phagocytosis.
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PMID:Enhanced phagocytic capacity. The biologic basis for the elevated histochemical nitroblue tetrazolium reaction. 415 97

This paper describes experiments conducted with membranous and soluble fractions obtained from Escherichia coli that had been grown on succinate, malate, or enriched glucose media. Oxidase and dehydrogenase activities were studied with the following substrates: nicotinamide adenine dinucleotide, reduced form (NADH), nicotinamide adenine dinucleotide phosphate, reduced form (NADPH), succinate, malate, isocitrate, glutamate, pyruvate, and alpha-ketoglutarate. Respiration was virtually insensitive to poisons that are commonly used to inhibit mitochondrial systems, namely, rotenone, antimycin, and azide. Succinate dehydrogenase and NADH, NADPH, and succinate oxidases were primarily membrane-bound whereas malate, isocitrate, and NADH dehydrogenases were predominantly soluble. It was observed that E. coli malate dehydrogenase could be assayed with the dye 2,6-dichlorophenol indophenol, but that porcine malate dehydrogenase activity could not be assayed, even in the presence of E. coli extracts. The characteristics of E. coli NADH dehydrogenase were shown to be markedly different from those of a mammalian enzyme. The enzyme activities for oxidation of Krebs cycle intermediates (malate, succinate, isocitrate) did not appear to be under coordinate genetic control.
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PMID:Respiration and protein synthesis in Escherichia coli membrane-envelope fragments. I. Oxidative activities with soluble substrates. 430 12

1. After conventional fractionation of rat liver homogenates in 0.88m-sucrose the mitochondrial fraction was subjected to short-term water lysis followed by separation of the resulting membrane preparations. 2. Phosphatidate formation was measured in all subcellular fractions and subfractions and was compared with the distribution of succinate dehydrogenase, monoamine oxidase, rotenone-insensitive NADH cytochrome c reductase, arylsulphatase, urate oxidase, arylesterase and glucose 6-phosphatase. 3. The results obtained indicated that mitochondria were capable of synthesizing phosphatidate, though this activity was only about one-third of the total homogenate activity. 4. Mitochondrial phosphatidate formation was located predominantly in the outer mitochondrial membrane. Although this membrane preparation was found to be significantly contaminated by the microsomal fraction, this contamination was estimated to account for not more than about 20% of the total phosphatidate formation observed in preparations of outer mitochondrial membrane.
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PMID:Phosphatidate biosynthesis in mitochondrial subfractions of rat liver. 430 22

Pyrrolnitrin at 10 mug/ml inhibited the growth of Saccharomyces cerevisiae, Penicillium atrovenetum, and P. oxalicum. The primary site of action of pyrrolnitrin on S. cerevisiae was the terminal electron transport system between succinate or reduced nicotinamide adenine dinucleotide (NADH) and coenzyme Q. At growth inhibitory concentrations, pyrrolnitrin inhibited endogenous and exogenous respiration immediately after its addition to the system. In mitochondrial preparations, the antibiotic inhibited succinate oxidase, NADH oxidase, succinate-cytochrome c reductase, NADH-cytochrome c reductase, and succinate-coenzyme Q(6) reductase. In addition, pyrrolnitrin inhibited the antimycin-insensitive reduction of dichlorophenolindophenol and of the tetrazolium dye 2,2'-di-p-nitrophenyl-(3,3'-dimethoxy-4,4'-bi-phenylene)5,5'-diphenylditetrazolium. The reduction of another tetrazolium dye, 2-p-iodophenyl-3-p-nitrophenyl-5-phenyltetrazolium chloride, that was antimycin-sensitive, was also inhibited by pyrrolnitrin. The antibiotic had no effect on the activity of cytochrome oxidase, and it did not appear to bind with flavine adenine dinucleotide, the coenzyme of succinic dehydrogenase. In whole cells of S. cerevisiae, pyrrolnitrin inhibited the incorporation of (14)C-glucose into nucleic acids and proteins. It also inhibited the incorporation of (14)C-uracil, (3)H-thymidine, and (14)C-amino acids into ribonucleic acid, deoxyribonucleic acid, and protein, respectively. The in vitro protein synthesis in Rhizoctonia solani and Escherichia coli was not affected by pyrrolnitrin. Pyrrolnitrin also inhibited the uptake of radioactive tracers, but there was no general damage to the cell membranes that would result in an increased leakage of cell metabolites. Apparently, pyrrolnitrin inhibits fungal growth by inhibiting the respiratory electron transport system.
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PMID:Mechanism of action of the antifungal antibiotic pyrrolnitrin. 431 80

1. Treatment of washed rat liver microsomes in a medium containing 0.12m-sucrose, 12.5mm-potassium chloride, 2.5mm-magnesium chloride and 25mm-tris-hydrochloric acid buffer, pH7.6, with 2m-lithium chloride at 5 degrees for 16hr. leads to the formation of membranes free of ribosomes and ribosomal subunits. 2. Confirmation of the absence of ribosomes from lithium chloride-prepared membranes was obtained by treatment of the membranes with sodium deoxycholate, followed by sucrose-density-gradient centrifugation, which showed the complete absence of ribosomes. 3. Treatment of membranes with phenol, followed by sucrose-density-gradient analysis of the isolated RNA, showed the presence of a small amount of 4s material. Repetition of the phenol extraction procedure in the presence of liver cell sap as a ribonuclease inhibitor again showed the presence of only 4s material. The 4s RNA was shown to be transfer RNA by the fact that it had the same capacity for accepting (14)C-labelled amino acids as isolated transfer RNA from rat liver pH5 enzyme. 4. Analysis showed that microsomes and membranes possessed similar glucose 6-phosphatase, NADH-2,6-dichlorophenol-indophenol reductase, NADH-neo-tetrazolium reductase, NADH-cytochrome c reductase and ribonuclease activities. 5. (3)H-labelled ribosomal RNA binds to membranes. However, isolation of the bound RNA by the phenol extraction procedure, followed by sucrose-density-gradient analysis, shows the RNA to be degraded to 7s material. Very little breakdown of (3)H-labelled ribosomal RNA bound to membranes occurs if the binding and isolation are carried out in the presence of liver cell sap.
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PMID:Preparation of ribosome-free membranes from rat liver microsomes by means of lithium chloride. 431 14

1. With reference to the post-operative dysfunction of the liver observed after halothane anaesthesia, the effects of the anaesthetic on some metabolic functions were studied in the isolated perfused rat liver. Oxygen uptake, glycolysis, gluconeogenesis and urea synthesis were affected by halothane at a concentration (2.5% of the gas phase) within the range used in clinical anaesthesia. 2. At this concentration of halothane uptake of oxygen was inhibited in livers from both fed and starved rats. 3. In livers from fed rats there was a 16-fold increase in lactate production. This was accompanied by a fivefold decrease in the tissue content of 2-oxoglutarate and a more than twofold decrease in citrate. The calculated [free NAD(+)]/[free NADH] ratio in both cytoplasm and mitochondria was lower in the halothane-exposed livers than in controls. 4. In livers of starved rats the rate of gluconeogenesis from lactate was decreased by halothane to 30% of the control rate. 5. Halothane inhibited gluconeogenesis from alanine and propionate to the same extent as from lactate, whereas glucose formation from dihydroxyacetone, glycerol, fructose and sorbitol was relatively unaffected. 6. During gluconeogenesis from 10mm-lactate the tissue content of ATP was decreased by 50%, glutamate by 50% and 2-oxoglutarate was decreased eightfold in the halothane-exposed livers. 7. Halothane decreased urea synthesis in the presence of 10mm-NH(4)Cl and 2mm-ornithine to 15% of the control rate. 8. The inhibitions of gluconeogenesis and urea synthesis were completely abolished within 15min of withdrawal of the anaesthetic. 9. The stimulation of uptake of oxygen brought about by the addition of lactate or precursors of urea was abolished by halothane. 10. Effects on gluconeogenesis similar to those of halothane occurred in livers exposed to the anaesthetic methoxyflurane, although normal rates were not restored on withdrawal of the drug. Other anaesthetic agents tested (ketamine-HCl and trichloroethylene) decreased gluconeogenesis to 66% of the control rate. 11. The inhibitory effects of halothane are consistent with an interference at the stage of the NADH dehydrogenase of the electron-transport chain.
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PMID:The effects of halothane (2-bromo-2-chloro-1,1,1-trifluoroethane) on glycolysis and biosynthetic processes of the isolated perfused rat liver. 434 8

Isopycnic equilibration and sedimentation rate studies of rat liver microsomes led previously to the assignment of microsomal constituents into group a1 (monoamine oxidase), group a2 (5'-nucleotidase, alkaline phosphodiesterase I, alkaline phosphatase and cholesterol), group a3 (galactosyltransferase), group b (NADH cytochrome c reductase, NADPH cytochrome c reductase, aminopyrine demethylase, cytochrome b(5) and P 450), and group c (glucose 6-phosphatase, esterase, nucleoside diphosphatase, beta-glucuronidase and glucuronyltransferase). Confirmation and extension of the assignment into groups has been obtained by studying the differential effect of the reagents digitonin, EDTA, and PPi. Digitonin specifically affected the equilibrium density only of the group a2 and (to a lesser extent) group a3, and not of groups b and c under conditions which preserved the structure-linked latency of nucleoside diphosphatase and galactosyltransferase. Within experimental error the rate of sedimentation of all microsomal constituents was unaffected. The morphological appearance under the electron microscope was indistinguishable from that of nondigitonin-treated microsomes, except that a few smooth membranes (< 10%) exhibited broken-looking profiles. Treatment of microsomes with EDTA or PPi detached a substantial part of RNA and released protein in excess over the amount accountable for by detachment of ribosome constituents. This detachment was confirmed by electron microscopy. EDTA and PPi decreased markedly the equilibrium density and the density dispersion of groups b and c, due mainly to the uncoating of rough elements. EDTA and PPi shifted slightly the distribution profiles of groups a towards lower densities, possibly as a result of the release of adsorbed proteins. The combination of EDTA and digitonin, used subsequently, rendered the average equilibrium density of group a2 higher than that of groups b and c. Dense subfractions were thus enriched in constituents of group a2 and showed mainly broken-looking vesicles under the electron microscope. The import of our results on the biochemical and enzymic properties of the subcellular components of the microsome fractions is discussed.
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PMID:Analytical study of microsomes and isolated subcellular membranes from rat liver. IV. Biochemical, physical, and morphological modifications of microsomal components induced by digitonin, EDTA, and pyrophosphate. 436 10

1. Starvation for 3 days produces a decrease in methaemoglobin-reductase and glutathione-reductase activities, but it does not alter the glucose 6-phosphate-dehydrogenase activity of the rat erythrocyte. 2. The feeding of a protein-free diet for 11 days causes greater changes in the first two enzymes and also a diminution of the third. Under this experimental condition slight decreases in protein and haemoglobin contents were noted. 3. The experimental animals did not show methaemoglobinaemia, probably because the activity of methaemoglobin diaphorase is preserved. 4. The GSH content was not affected but the stability of the tripeptide in the presence of an oxidizing agent was diminished.
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PMID:Studies on the oxidation-reduction systems of the erythrocyte. 437 99


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