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:1.7.1.2 (nitrate reductase)
3,861 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cell extract from a strain of Propionibacterium acidi-propionici with high nitrate reductase (NaR) activity catalyzed nitrate reduction with glycerol phosphate, NADH, or lactate. The reaction was inhibited partially by fumarate or oxygen. NaR linked to methyl viologen was found mostly in particulate fractions. It was solubilized by treatment with Emulgen 810 and purified 46-fold by DEAE-cellulose, Sepharose 4B, and triple DEAE-Sephadex chromatographies in the presence of the detergent. It was rather labile but was stabilized by glycerol. The molecular weight was estimated to be 230,000 by Sepharose 4B gel filtration and the isoelectric point was pH 5.0-5.5. The pH optimum was at 6.5-7.5 and Km for nitrate was 0.1 mM. As electron donors, methyl and benzyl viologen were utilized well but FAD and FMN were fairly ineffective. Chlorate was an active acceptor as well as nitrate. Azide, cyanide, and thiocyanate inhibited NaR. On adding 1 mM tungstate to the growing medium, the NaR level in grown cells was lowered; addition of 0.01 mM molybdate restored the activity partially. NaR is suggested to be a molybdo-protein, similar to this enzyme from other bacteria.
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PMID:A study on nitrate reductase from Propionibacterium acidi-propionici. 62 3

Active transport of amino acids by membrane vesicles from Escherichia coli, grown anaerobically on glucose in the presence of nitrate, can be energized under anaerobic conditions by electron transfer in the nitrate respiration system with formate as electron donor and nitrate as acceptor. A high rate of amino acid transport is also obtained under anaerobic conditions by electron transfer from formate to the nitrate analogue chlorate or to the membrane-impermeable electron acceptor ferricyanide. Electron transfer from formate to nitrate results in the generation of an electrical potential as is indicated by the uptake of the lipophilic cation triphenylmethylphosphonium. Ferricyanide accpets electrons from at least two sites of the nitrate respiration system. One of these sites appears to be nitrate reductase, because cytochrome b, reduced by formate, is completely reoxidized by ferricyanide and glutamate transport energized by formate plus ferricyanide and formate plus nitrate are affected by the same electron transfer inhibitors. A second site of electron transfer to ferricyanide appears to be located prior to nitrate reductase in the nitrate respiration system, since formate is oxidized at a higher rate in the presence of ferricyanide than with nitrate while formate/ferricyanide energizes transport of amino acids at a lower rate than formate/nitrate. Moreover, electron transfer inhibitors block electron transfer from formate to nitrate to a significantly higher extent than from formate to ferricyanide. The effects of irradiation of the membrane vesicles with near ultra-violet light suggest that quinones play an essential role in the electron transfer from formate to nitrate or ferricyanide. Irradiation blocks completely formate-dependent nitrate and ferricyanide reduction and active transport driven by formate/nitrate and formate/ferricyanide, but has hardly any effect on the activity of formate dehydrogenase and on ascorbate/phenazine methosulphate/oxygen-driven transport. Similar effects of ferricyanide have been observed in membrane vesicles from E. coli, grown anaerobically in the presence of fumarate. In these membrane vesicles a high rate of lactose and triphenylmethylphosphonium uptake under anaerobic conditions is obtained by electron transfer from glycerol 1-phosphate to fumarate and also to ferricyanide and evidence has been presented for the involvement of cytochromes in these electron transfers.
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PMID:Active transport by membrane vesicles from anaerobically grown Escherichia coli energized by electron transfer to ferricyanide and chlorate. 79 48

Yersinia was investigated in 50 skin samples of chicken carcasses from retail shops and 65 samples of balanced food for domestic fowl. Enrichments were performed in saline phosphate buffer 0.067 M, pH 7.6 and post-enriched in 0.5% KOH. Subcultures were performed in Salmonella-Shigella agar and MacConkey agar. Isolates were identified through biochemical, serological and lysotyping methods. The following biovar (B), serovar (O) and phagovar (Lis) were isolated from chickens: Y. enterocolitica (five strains) B:1:O:6,47;Lis Xz; B:1;O:6:Lis Xz; B:1:O:5,Lis Xz; Y.intermedia (two strains) B:1;O:52;Lis Xz; B:1;O:52,53,54;Lis Xz (NRA, nitrate reductase type A); Y. frederiksenii (two strains) O:10,K1,25,35,38,46:Lis Xz; (citrate): O:10,K1,25,35,38,46:Lis Xz (ONPG-: citrate +); Y. kristensenii (one strain) does not agglutinate; Lis Xo. Yersinia were not isolated from balanced food for domestic fowl. Virulence tests (calcium dependency and autoagglutination at 37 degrees C) were negative in all instances. It is concluded from this study that Yersinia isolated from chickens are without pathogenic importance.
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PMID:[Bacteria of the genus Yersinia in chickens for human consumption and in balanced bird food]. 181 65

Electron paramagnetic resonance spectra obtained during turnover of the Mo center of NADH:nitrate reductase at pH 8 were comprised of two Mo(V) species, signal A (g1 = 1.996, g2 = 1.969, g3 = 1.967, A1H = 1.25 mT, A2H = 1.18 mT, and A3H = 1.63 mT) and signal B (g1 = 1.996, g2 = 1.969, and g3 = 1.967), the former exhibiting superhyperfine interaction due to strong coupling with a single, exchangeable proton. Binding of halides and nitrite to the Mo center increased the proportion of signal A whereas phosphate had no effect on the EPR line shape. Halides decreased and phosphate increased the rates of enzyme activities involving the Mo center (NADH:nitrate reductase and reduced methyl viologen:nitrate reductase), but neither had any effect on activities involving FAD (NADH:ferricyanide reductase) or heme (NADH:cytochrome c reductase), indicating specific binding of halides to the Mo center. Halides were found to be weak, mixed competitive-noncompetitive inhibitors (Cl- KI = 39 mM, mu = 0.2 M, pH 8) of nitrate reductase forming a catalytically inactive ternary halide-nitrate-enzyme complex. Inhibition patterns changed from nearly noncompetitive (F-) to nearly competitive (I-). The weakening of nitrate binding due to halide binding correlated with increased halide electronegativity rather than ionic radius. In contrast, phosphate (Kd = 7.4 mM, mu = 0.2 M, pH 8) and arsenate were determined to be nonessential activators, characterized by a constant value of (Vmax/Km)app, increasing nitrate reductase activity by weakening nitrate binding without affecting the stability of the transition state. Phosphate had no effect on product inhibition by nitrite (KI = 0.33 mM) or the oxidation-reduction midpoint potentials of the Mo center.
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PMID:EPR and kinetic analysis of the interaction of halides and phosphate with nitrate reductase. 255 63

Molybdenum cofactor (mocofactor) is extracted efficiently, free of impurities and in high concentrations, by acid treatment of xanthine oxidase and subsequent incubation of the precipitate with phosphate buffer containing EDTA, molybdate and oxygen. It is suggested that cofactor is bound to the enzyme via hydrophobic forces as well as via an oxygen-sensitive mechanism. Upon extraction, the capability to complement the apo nitrate reductase of Neurospora crassa nit-1 can be conserved only in the total absence of oxygen. Cysteine and glutathione were shown to protect efficiently free mocofactor from oxidation. Two species of active mocofactor, probably a molybdoform and a demolybdoform, could be separated by means of reversed-phase HPLC with a mobile phase of 5 mM sodium citrate at a pH of 6.5. The mode of interaction between either of these species with thiol reagents is discussed.
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PMID:Extraction and purification of molybdenum cofactor from milk xanthine oxidase. 369 96

The molybdopterin cofactor from the formate dehydrogenase of Methanobacterium formicicum was studied. The cofactor was released by guanidine denaturation of homogeneous enzyme, which also released greater than 80% of the molybdenum present in the enzyme. The anoxically isolated cofactor was nonfluorescent, but after exposure to air it fluoresced with spectra similar to those of described molybdopterin cofactors. Aerobic release from acid-denatured formate dehydrogenase in the presence of I2 and potassium iodide produced a mixture of fluorescent products. Alkaline permanganate oxidation of the mixture yielded pterin-6-carboxylic acid as the only detectable fluorescent product. The results showed that the cofactor from formate dehydrogenase contained a pterin nucleus with a 6-alkyl side chain of unknown structure. Covalently bound phosphate was also present. The isolated cofactor was unable to complement the cofactor-deficient nitrate reductase of the Neurospora crassa nit-1 mutant.
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PMID:Molybdopterin cofactor from Methanobacterium formicicum formate dehydrogenase. 370 Mar 35

The oxidation-reduction midpoint potential for the heme prosthetic group present in assimilatory nitrate reductase from Chlorella vulgaris has been determined by optical potentiometric titrations in the presence of dye mediators. At pH 7, the midpoint potential was determined to be -160 mV and corresponds to a reversible n = 1 redox process. The midpoint potential was unaltered by the use of NADH as reductant, unaffected by the presence of NAD+, cytochrome c, phosphate, cyanide, or alkaline pH. In addition, the redox potential of the heme was independent of modifications to the enzyme such as substitution of the molybdenum center with tungsten, or cleavage and separation of the enzyme into its flavin and heme/molybdenum domains. In contrast, the midpoint potential increased on decreasing the pH yielding a pH dependence of approximately 20 mV/pH unit within the range 5.5 to 7, suggesting the presence of a single, redox-associated, ionizable functional group on the protein with pKox = 5.8 and pKred = 6.1. At pH 7 and within the range 12 to 38 degrees C, the midpoint potential of the heme decreased by approximately 1 mV/degree. Values for delta S0 and delta H0 were calculated to be -25.6 e.u. and -4.0 kcal/mol.
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PMID:Thermodynamic properties of the heme prosthetic group in assimilatory nitrate reductase. 370 Mar 73

Initial velocity studies of Chlorella nitrate reductase showed that increased ionic strength stimulated NADH:nitrate reductase activity by increasing both Vmax and Km for nitrate. Examination of the effect of ionic strength on the various partial activities of nitrate reductase revealed that while NADH:ferricyanide and reduced methyl viologen:nitrate reductase activities were unaffected by ionic strength, NADH:cytochrome c and reduced flavin:nitrate reductase activities were inhibited and stimulated by increased ionic strength, respectively. Comparison of the rates for the partial activities indicated electron transfer from heme to molybdenum to be the rate-limiting step in enzyme turnover. The pH optimum for NADH:nitrate reductase activity was found to be 7.9 while values for the partial activities ranged from 5.5 to 8.1. Phosphate was found to stimulate both NADH:nitrate and reduced methyl viologen:nitrate reductase activities indicating the molybdenum center as the site of interaction.
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PMID:Assimilatory nitrate reductase from Chlorella. Effect of ionic strength and pH on catalytic activity. 377 27

During anaerobic growth, Escherichia coli can reduce phosphomolybdate. The reduction can also be carried out by washed cells suspended in buffer at pH 5.7. Phosphate, molybdate, glucose, cells, and anaerobic conditions are required. Reduction is inhibited by 200 microM chromate, 290 microM nitrite, 10 mM tungstate, or 20 mM cysteine. Wild-type (chl+) cells are inhibited by addition of 200 microM nitrate, but chlA, chlB, and chlE mutants are not. The inhibition of chl+ cells results from reduction of nitrate to nitrite. This nitrate reduction is not catalyzed by nitrate reductase. Wild-type cells are more sensitive than chl mutants to inhibition by nitrite and cysteine but more resistant to chromate. Pregrowth of chlD cells in 1 mM Na2MoO4 increases their sensitivity to nitrite and cysteine, and pregrowth of chl+ cells in 1 mM Na2MoO4 increases their resistance to these agents. Assays of biotin sulfoxide reductase show that the tightness of the chlD block depends on growth conditions; chlD cells grown aerobically in tryptone broth make about 50% as much active enzyme as chl+ cells, whereas chlD cells grown anaerobically with tryptone plus glucose make less than 10%. The effect of anaerobic pregrowth on the inhibition of molybdate reduction by added nitrate indicates that in vivo nitrate reduction responds to growth conditions in the same manner as biotin sulfoxide reductase does.
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PMID:Molybdate reduction by Escherichia coli K-12 and its chl mutants. 388 54

Nitrate reductase (EC 1.6.6.1) from Chlorella vulgaris, a flavin-cytochrome-molybdenum enzyme, catalyses two types of partial reactions: reduction of exogenous cytochrome c by NADH and reduction of nitrate to nitrite by reduced methyl viologen (reduced 1,1'-dimethyl-4,4'-dipyridine dichloride). Ferrate, an analogue of orthophosphate acting on the phosphate-binding region of the enzymes, abolishes the NADH-nitrate reductase as well as the NADH-cytochrome c activities. In addition, the ability of NADH to reduce the endogenous cytochrome b component of the enzyme is also impaired. The reduction of nitrate by reduced methyl viologen is only partially affected. The results indicate that the ferrate primarily disrupts the NADH site.
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PMID:Inactivation of the NADH-dependent activities of nitrate reductase by ferrate. 409 Aug 56


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