Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:Q8NEX9 (reductase)
 26,410 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Ribonucleoside diphosphate reductase from Escherichia coli consists of a 1/1 complex of two nonidentical subunits called proteins B1 and B2. The enzyme reduces the four common ribonucleoside diphosphates to the corresponding deoxyribonucleotides and is allosterically regulated by nucleoside triphosphates which influence its substrate specificity as well as its overall activity. The B1 subunit contains binding sites for the effectors while B2 contains iron and an organic free radical essential for catalytic activity. We now establish that only protein B1 binds substrates. Competition experiments support the presence of two identical substrate binding sites, distinct from the effector binding sites. The catalytic site of the enzyme thus is formed from both the B1 and B2 subunits. Dissociation constants for substrates ranged from 2 X 10(-5) to about 10(-3) M. In all cases effectors decreased these constants in agreement with their influence on the substrate specificity of ribonucleotide reductase, but did not induce cooperative effects. The increase in binding was pronounced at 20 degrees but only marginal at 0 degrees. Arrhenius plots of the influence of temperature on the catalytic activity of the enzyme showed sharp breaks at 12 degrees. The temperature effects can be interpreted as a conformational change occurring in the structure of protein B1 at the critical temperature.
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PMID:Binding of substrates to Escherichia coli ribonucleotide reductase. 77 72

Comparison of various chloroplast-type ferredoxin sequences, chemical and enzymic modifications, reconstitution experiments, and fluorescence measurement of chloroplast-type ferredoxins have led to the following conclusions. 1. Tyrosine, histidine, and tryptophan residues are not directly involved in the oxidation-reduction mechanism of ferredoxins. The four indispensible cysteine residues in spinach ferredoxin which constitutes a part of the iron-sulfur cluster are located at residues 39, 44. 47 and 77. Two out of six cysteine residues in Spirulina ferredoxin could be easily modified with vinylpyridine without the loss of reconstitutive ability i.e. the apoferredoxin could be converted to the holoform by the addition of iron and sulfide. 2. Spinach ferredoxin was digested with carboxypeptidase A and the terminal alanine could be removed without loss of the spectral properties of native ferredoxin. However, the removal of the terminal three residues gave rise to the loss of reconstitutive ability. 3. The amino groups of spinach ferredoxin were modified by acetic anhydride and four residues were acetylated. The acetylated preparation of ferredoxin had an unique spectrum. Upon the addition of high concentration of ions the spectrum of this derivative resembled the spectrum of native ferredoxin. Acetylferredoxin did not combine with ferredoxin-NADP reductase, but upon the addition of moderate concentrations of cations, it did bind to this enzyme.
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PMID:Structure and function of chloroplast-type ferredoxins. 78 73

Ribonucleoside diphosphate reductase determined by bacteriophage T4 consists of a tight complex (alpha2beta2) of the polypeptide chains alpha (Mr = 80,000 to 85,000) and beta (Mr = 35,000). The alpha2 dimer (= protein B1) was purified from Escherichia coli B infected with T4 mutant nrdB55 (Yeh, Y.C., and Tessman, I. (1972) Virology 47, 767-772) which carries an amber mutation in the gene coding for the beta polypeptide chain. Protein B1 contained binding sites for dATP, an allosteric effector of the reductase. The beta2 dimer (= protein B2) was purified by selective desorption with 1 M guanidine HCl from a dATP-Sepharose affinity column containing adsorbed native T4 ribonucleotide reductase. Protein B2, isolated this way, was enzymatically inactive due to partial loss of its iron but it could be reactivated by treatment with ferrous iron. Active protein B2 contained two atoms of non-heme iron per molecule and exhibited the optical and electron spin resonance spectra previously demonstrated in the native enzyme. The T4-induced proteins B1 and B2 were unable to reduce ribonucleotides when assayed separately but were active in combination. The proteins did not form catalytically functional hybrids with proteins B1 and B2 of Escherichia coli ribonucleotide reductase, neither did they cross-react immunologically with the latter. 5-Hydroxymethyl-dCTP, at concentrations above 10 muM, was a positive allosteric effector of T4 ribonucleotide reductase promoting the reduction of the pyrimidine ribonucleotides CDP and UDP. The nucleotide had little effect on E. coli ribonucleotide reductase.
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PMID:Ribonucleoside diphosphate reductase induced by bacteriophage T4. III. Isolation and characterization of proteins B1 and B2. 80 36

An iron-sulfur protein has been isolated from chick kidney mitochondria and purified (200-fold as determined enzymatically by its NADPH-cytochrome c reductase activity in the presence of adrenodoxin reductase) on DEAE-cellulose and gel filtration on Sephadex G-100. The purified protein showed an absorption peak at 411 nm with a shoulder at 460 nm. The electron paramagnetic resonance spectrum was typical of a ferredoxin-type iron-sulfur protein with g values: gx=gy-1.94 and gz=2.02. The molecular weight was estimated by gel filtration to be 12,500. When tested against anti-adrenodoxin gamma-globulin, the protein showed a precipitin line that fused completely with that of adrenodoxin. Based on these findings it is concluded that this protein is an iron-sulfur protein quite similar to adrenal ferredoxin. In the presence of adrenoxodin reductase, NADPH, and carbon monoxide, the purified renal ferredoxin was found to be active in the reduction of cytochrome P-450 solubilized from chick kidney mitochondria. It was also effective in the reconstituted 25-hydroxyvitamin D3-1alpha-hydroxylase composed of the cytochrome P-450 from rachitic chick kidneys and adrenodoxin reductase. A ferredoxin reductase isolated from chick kidney mitochondria could replace adrenodoxin reductase in the reconstituted system. These results strongly support a previous conclusion that the kidney mitochondrial 25-hydroxyvitamin D3-1alpha-hydroxylation system consists of a renal ferredoxin reductase (presumably a flavoprotein), renal ferredoxin, and cytochrome P-450.
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PMID:Isolation of chick renal mitochondrial ferredoxin active in the 25-hydroxyvitamin D3-1alpha-hydroxylase system. 81 34

The reaction mechanism of adenosine 5'-phosphosulfate (APS) reductase (EC 1.8.99.2) from Thiobacillus thioparus was studied using difference spectrum and stopped-flow techniques. The enzyme-bound FAD was rapidly reduced by sulfite with a first order rate constant of 97.1 s-1. The addition of AMP induced further spectral changes in the reduced enzyme which were consistent with the oxidation of FADH2 to the red (anionic) semiquinone FADH-) and the concomitant reduction of nonheme iron to the ferrous state. Superoxide dismutase (EC 1.15.1.1) or anaerobiosis inhibited the reduction of cytochrome c by the enzyme only to the extent of 25-35%, indicating the existence of a direct reduction of cytochrome c by the enzyme without involving O2-. the activity of enzyme with cytochrome c was inhibited by increasing the potassium phosphate concentration, the inhibition being more pronounced with horse heart cytochrome c than with Candida krusei cytochrome c.
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PMID:A study on the reaction mechanism of adenosine 5'-phosphosulfate reductase from Thiobacillus thioparus, an iron-sulfur flavoprotein. 83 49

Rubredoxin, one of the three protein components of the epoxidation/hydroxylation system of Pseudomonas oleovorans was immobilized by attachment to CNBr-activated agarose (Sepharose 4B). Since this represents the first reported example of the preparation of a water-insoluble derivative of an enzyme of this type, the electron transfer and physical properties of the conjugate were examined in order to allow comparison with those of the soluble enzyme. Immobilized rubredoxin exhibits all of the major spectral properties of the soluble enzyme above 300 nm, but some distortion in the 280 nm abosrbance band was observed. The immobilized enzyme accepts electrons from dithionite or form NADPH in the presence of spinach ferredoxin-NADP reductase, and upon reduction the visible absorbance is bleached. Immobilized rubredoxin mediates the reduction of cytochrome c in the presence of NADPH and spinach reductase, although it is less efficient in this role than soluble rubredoxin. The oxidation-reduction potential of immobilized rubredoxin was determined and found to be similar to that of the soluble enzyme. In the presence of 2.5 m guanidine HCL, the immobilized enzyme is considerably more stable than soluble rubredoxin toward denaturation. After anaerobic reduction, iron was readily removed from immobilized rubredoxin by washing in 0.5 m Tris base, PH 9.5 containing 0.07 M mercaptoethanol, and the resulting immobilized apoenzyme could then be reconstituted to give back a conjugate with the original iron content, as judged from its absorbance at 497 NM. Reptition of the entire reduction-dissociation-reconstitution cycle gave the same results as were obtained after the initial reconstitution.
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PMID:Preparation and properties of immobilized rubredoxin. 84 34

The study of the participation of metals in evolution of oxidation-reduction processes is subdivided into two periods. During the first of them, from 1897 to 1937, the significance of manganese, iron, titanium, molybdenum, vanadium and copper in most important processes of metabolism was discovered. The second period, from 1937 to 1977, was devoted to the study of the role of metals in individual representatives of oxidoreductases and their evolution during transition of organisms from anaerobiosis to aerobiosis. In this evolution of special importance were bimetallic enzymes, such as nitrogenase, some nitrate reductases and hydrogenases, carbon dioxide reductase, xanthine oxidase, cytochrome oxidase. Owing to their ability to accomplish conjugated oxidation-reduction reactions, these oxidoreductases were transitional to still more complicated polymetallic systems with whose participation the electron transfer chains in subcellular structures were formed.
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PMID:[Participation of polyvalent metals in the evolution of oxidoreductases]. 91 1

Studies on the effects of inhibitors of the nitrate-reducing activity of Veillonella alcalescens extracts suggest the participation of a naphthoquinone, a b-type cytochrome, and non-heme iron in electron transport to nitrate. A nitrate-reductase-deficient mutant displayed a longer doubling time and a decreased molar growth yield on nitrate media. This mutant was phenotypically restored by the addition of molybdate to the growth medium, giving evidence for the functioning of molybdenum in the nitrate-reductase enzyme of V. alcalescens.
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PMID:Nitrate-reductase electron-transport cofactors in Veillonella alcalescens. 92 6

A crystalline NADPH-adrenodoxin reductase was obtained from bovine adrenocortical mitochondria and its properties were investigated. Its molecular weights and isoelectric point were estimated to be 51 000 and 5.4, respectively. Amino acid and sugar contents and the interaction between the apo-reductase and flavin of NADPH-adrenodoxin reductase were investigated. Formation of a complex of bovine NADPH-adrenodoxin reductase with adrenodoxin, its apoadrenodoxin, or other non-heme iron proteins caused quenching of fluorescence of the tryptophanyl residue and bound FAD of the NADPH-adrenodoxin reductase. The results obatined suggest that adrenodoxin and apoadrenodoxin bind functionally to a site close to the tryptophanyl residue and the bound FAD of the reductase. The circular dichroism spectrum of oxidized NADPH-adrenodoxin reductase was measured in the ultraviolet and visible regions. This spectrum showed negative absorption in the visible region and was not appreciably influenced in either the ultraviolet or visible region by formation of a complex with adrenodoxin or apoadrenodoxin.
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PMID:Properties of crystalline reduced nicotinamide adenine dinucleotide phosphate-adrenodoxin reductase from bovine adrenocortical mitochonria. I. Physicochemical properties of holo- and apo-NADPH-adrenodoxin reductase and interaction between non-heme iron proteins and the reductase. 98 53

A new catalitic activity of soluble succinate dehydrogenase, i.e. the reduction of low (20-200 muM) concentration of ferricyanide in the presence of succinate is described. The apparent Km value for the acceptor is about 200 muM. The turnover numbers of the enzyme measured in this reaction, with PMS as an electron acceptor and in the system reconstituted from soluble enzyme and alkali-treated submitochondrial particles (succinate oxidase) are found to be almost the same. The new succinate. ferricyanide reductase activity is very sensitive to oxygen, high (3 mM) ferricyanide concentration and mercaptide-forming agents. When the enzyme is stored under aerobic conditions the loss of this activity occurs according to the first-order kinetics with the same rate constants as the reconstitutive activity decreases. The rate constants both for ferricyanide reductase and reconstitution decay do not depend on pH within the range of 6,5--7,5 (k = 8.10(-2) min-1) and increase dramatically at pH 8,5 (K = 4.10(-1) MIN-1). When these two activities are lost after oxygen exposure the PMS-reductase fall down to about 50% of its original activity. The new ferricyanide reductase is found only in the soluble preparation of the enzyme succinate: cytochrome c reductase, succinate dehydrogenase of submitochondrial particles and reconstituted succinate oxidase do not interact with low concentrations of ferricyanide. The treatment of the enzyme after inactivation by oxygen exposure with sulfide ion--iron--mercaptoethanol mixture followed by Sephadex filtration completely restores the original reconstitutive, ferricyanide and PMS reductase activities. The hypothesis is suggested that succinate dehydrogenase contains at least two red-ox centers reacting with electron acceptors. The first one is located in hydrophylic environment (mitochondrial matrix) being accessible for high concentrations of ferricyanide. The second one (iron--sulfur complex, Hipip-type) is responsible for ferricyanide reductase activity described, being located intramembraneously and involved in the electron transfer between dehydrogenase and the rest of the respiratory chain.
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PMID:[Kinetic and structural characteristics of succinate dehydrogenase components reacting with natural and artificial electron acceptors]. 99 75


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