UNIPROT:Q8NEX9 - FACTA Search

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Query: UNIPROT:Q8NEX9 (reductase)
26,410 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

[14C]Cortisol was injected iv into three subjects during a control period and while receiving metyrapone. The plasma kinetics of the tracer cortisol and the patterns of its urinary metabolites were measured. Metyrapone caused an increase in the volume of distribution of cortisol (34%) and in the MCR (75%); the half-life was decreased by 25%. There were marked changes in the urinary metabolite pattern: 3 alpha,11 beta,17,21-tetrahydroxy-5 alpha-pregnan-20-one, 3 alpha,17,21-trihydroxy-pregnane-11,20-dione(THE), pregnane-3 alpha,11 beta,17,20 alpha,21-pentol, plus pregnane-3 alpha,11 beta,17,20 beta-21-pentol (cortol), and 3 alpha,17,20 alpha,21-tetrahydroxypregnan-11-one (cortolone) all decreased by an average of 62%, 44%, 38%, 45%, and 25% respectively. In contrast, there was an increase of 296% in 3 alpha,17,20 beta,21-tetrahydroxypregnan-11-one (beta-cortolone). To account for these effects it is postulated that metyrapone has the following extraadrenal actions: 1) it inhibits the back reduction of cortisone to cortisol and 2) it stimulates the 20-ketosteroid reductase that converts THE to beta-cortolone.
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PMID:Extraadrenal effects of metyrapone in man. 26 28

The study of the nickel enzyme methyl-coenzyme M reductase from methanogenic bacteria has been hampered until now by the fact that upon cell rupture the activity of the enzyme always dropped to at best only a few percent of its in vivo activity. We describe here that when Methanobacterium thermoautotrophicum cells were preincubated with 100% H2 before disintegration methyl-coenzyme M reductase activity stayed high. The cell extracts with a specific activity of 2 U/mg protein exhibited two nickel-derived EPR signals, designated MCR-red1 and MCR-red2, previously only observed in intact cells. The enzyme was purified 10-fold to a specific activity of 20 U/mg in the presence of methyl-coenzyme M, which stabilized both the activity and the EPR signal MCR-red1. The enzyme preparation displayed an UV/Vis spectrum with an absorption maximum at 386 nm and a shoulder at 420 nm. Upon inactivation of the enzyme with O2 or CHCl3, the maximum at 386 nm and the EPR signals MCR-red1 and MCR-red2 disappeared.
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PMID:Methyl-coenzyme M reductase preparations with high specific activity from H2-preincubated cells of Methanobacterium thermoautotrophicum. 165 49

Purified S-methyl-coenzyme M reductase (methylreductase) exhibits a very low fraction of its in vivo activity, suggesting either enzyme inactivation during cell lysis and chromatographic purification or the lack of an activating component in assay mixtures. Evidence that all methylreductase molecules in the purified protein can catalyze slow substrate turnover is found in a study of turnover-dependent in vitro incorporation of radiolabeled HS-CoM at the enzyme active site (Hartzell, P. L., Donnelly, M. I., and Wolfe, R. S. (1987) J. Biol. Chem. 262, 5581-5586). We have conducted active site titrations of purified methylreductase and of a highly active partially purified preparation (Rospert, S., Bocher, R., Albracht, S. P. J., and Thauer, R. K. (1991) FEBS Lett. 291, 371-375) using the reversible competitive inhibitor bromopropanesulfonate (K(i) = 0.05 microM). Curve fitting the data based on an equilibrium binding model shows that 0.1-1.4% of purified methylreductase has functional inhibitor binding sites while up to 25% of a highly active preparation binds the inhibitor. An EPR titration of highly active methylreductase with this inhibitor is consistent with this result, showing that the MCR-red1 and -red2 EPR signals (Albracht, S. P. J., Ankel-Fuchs, D., Bocher, R., Ellermann, J., Moll, J., van der Zwann, J. W., and Thauer, R. K. (1988) Biochim. Biophys. Acta 955, 86-102) are titrated in parallel with this active fraction. Attempts to observe turnover-dependent uptake of radiolabel from [thio-35S]2-methylthioethane-sulfonate by methylreductase were unsuccessful. These results suggest that the low activity of purified methylreductase is due primarily to low percentages of catalytically competent enzyme.
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PMID:Nature of the low activity of S-methyl-coenzyme M reductase as determined by active site titrations. 839 4

The administration of exogenous testosterone (T) to eugonadal men causes suppression of gonadotropin secretion and thus of spermatogenesis. This is currently being investigated as a possible method of hormonal male contraceptive, but complete suppression of spermatogenesis to azoospermia is induced in only 50-70% of Caucasian men; the remainder maintain a low rate of spermatogenesis. The basis for this polymorphism in response is unclear. The enzyme 5 alpha-reductase (5 alpha R) converts T to dibydrotestosterone (DHT) and is important in determining the magnitude of the androgen stimulus in some tissues. We investigated whether the maintenance of spermatogenesis in men remaining oligozoospermic while receiving suppressive doses of T is associated with evidence of increased (TE), im, weekly in a clinical trial of hormonal male contraception. The MCR of T (MCRT) and the conversion ratio of T to DHT (CRT-DHT) were measured by infusion of [3H]T, plasma levels of DHT and androstanediol glucuronide (AdiolG) were measured by RIA, and 24-h urinary steroid metabolites were measured by capillary column gas chromatography. Sperm density decreased in all men; 18 achieved azoospermia by 20 weeks of treatment, and the remainder had a mean sperm density of 2.0 +/- 0.8 x and MCRT, but with no differences between azoospermic and oligozoospermic responders. There were no differences in CRT-DHT plasma DHT, or AdiolG before treatment, but after 16 weeks, CRT-DHT had increased in the oligozoospermic responders, but not in the azoospermic responders. TE treatment increased plasma DHT and AdiolG levels in both groups, but the increases in both 5 alpha R metabolites were significantly greater in the oligozoospermic responders. Urinary excretion of etiocholanolone and androsterone was increased after 16 weeks of TE treatment, but did not differ between the two groups, andetiocholanolone/androsterone ratios did not differ greatly from unity. There was no change in urinary excretion of tetrahydrocortisol, allo-tetrahydrocortisol, or cortisone after 16 weeks of TE treatment in either group. These results suggest that after TE administration there is a selective increase in 5 alpha R activity in those men who remain oligozoospermic, but not in those becoming azoospermic. This difference in the androgenic milieu may underlie the incomplete suppression in the oligozoospermic responders, in whom a low rate of spermatogenesis is maintained despite the apparent absence of gonadotropins. In the UK, physicians recruited 33 healthy men aged 21-41 to a clinical trial of weekly intramuscular injections of 200 mg testosterone enanthate (TE) (Testoviron) for up to 18 months. 18 (55%) of the men achieved azoospermia after 20 weeks of treatment. The sperm density in the remaining 15 men (45%) stabilized at a mean density of 2(+or- 0.8) million/ml and stayed oligozoospermic for the remainder of the clinical trial. Plasma samples taken immediately before and 1, 2, 4, and 7 days after the 1st and 16th TE injections and after 2, 4, 8, and 12 weeks of treatment allowed the researchers to compare the activity of 5alpha-reductase (5alphaR). 5alphaR converts T to the more potent androgen dihydrotestosterone (DHT) and plays a key role in determining the magnitude of the androgen stimulus in some tissues. Exogenous testosterone (T) increased plasma T levels and the mean conversion rate of T (MCRT), but there were no differences between azoospermic and oligozoospermic men. Before treatment there were no differences in plasma DHT and androstanediol glucuronide (AdiolG) levels between the two groups. After 16 weeks, the conversion ratio of T to DHT (CRT-DHT) increased in oligozoospermic men (3.1% vs. 4%; p 0.05) but not in azoospermic men, whereas before treatment it was essentially the same in both groups. Exogenous T significantly increased both plasma DHT and AdiolG levels in both groups, but the increases in both these 5alphaR metabolites were significantly higher in oligozoospermic men than azoospermic men (p 0.02 and 0.01, respectively). (Abstract Truncated)
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PMID:Comparison between testosterone enanthate-induced azoospermia and oligozoospermia in a male contraceptive study. III. Higher 5 alpha-reductase activity in oligozoospermic men administered supraphysiological doses of testosterone. 877 48

The nickel porphinoid, coenzyme F430, is the prosthetic group of methyl-coenzyme M reductase. The active form of the enzyme exhibits Ni-EPR signals designated as MCR-red1 and MCR-red2. The inactive form of the enzyme is either EPR silent or it exhibits a distinct Ni-EPR signal designated MCR-ox1. Evidence is presented here that the MCR-ox1 form of the enzyme can be converted in vitro to the MCR-red1 form by reduction with titanium(III) citrate at pH 9. During conversion, the specific activity increases with increasing MCR-red1 spin concentration from 2 U/mg to approximately 100 U/mg at spin concentrations higher than 80%. The reduced methyl-coenzyme-M reductase shows an ultraviolet/visible spectrum characteristic for coenzyme F430 in the Ni(I) oxidation state, with maxima at 386 nm and at 750 nm. The results indicate that methyl-coenzyme-M reductase is activated when the enzyme-bound coenzyme F430 is reduced to the Ni(I) oxidation state. The experiments were performed with purified methyl-coenzyme-M reductase isoenzyme I of Methanobacterium thermoautotrophicum (strain Marburg).
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PMID:Purified methyl-coenzyme-M reductase is activated when the enzyme-bound coenzyme F430 is reduced to the nickel(I) oxidation state by titanium(III) citrate. 903 Jul 28

The interconversion of hormonally active cortisol (F) and inactive cortisone (E) is catalyzed by two isozymes of 11beta-hydroxysteroid dehydrogenase (11betaHSD), an oxo-reductase converting E to F (11betaHSD1) and a dehydrogenase (11betaHSD2) converting F to E. 11betaHSD1 is important in mediating glucocorticoid-regulated glucose homeostasis and regional adipocyte differentiation. Earlier studies conducted with GH-deficient subjects treated with replacement GH suggested that GH may modulate 11betaHSD1 activity. In 7 acromegalic subjects withdrawing from medical therapy (Sandostatin-LAR; 20-40 mg/month for at least 12 months), GH rose from 7.1 +/- 1.5 to 17.5 +/- 4.3 mU/L (mean +/- SE), and insulin-like growth factor I (IGF-I) rose from 43.0 +/- 8.8 to 82.1 +/- 13.7 nmol/L (both P < 0.05) 4 months after treatment. There was a significant alteration in the normal set-point of F to E interconversion toward E. The fall in the urinary tetrahydrocortisols/tetrahydocortisone ratio (THF+allo-THF/THE; 0.82 +/- 0.06 to 0.60 +/- 0.06; P < 0.02) but unaltered urinary free F/urinary free E ratio (a marker for 11betaHSD2 activity) suggested that this was due to inhibition of 11betaHSD1 activity. An inverse correlation between GH and the THF+allo-THF/THE ratio was observed (r = -0.422; P < 0.05). Conversely, in 12 acromegalic patients treated by transsphenoidal surgery (GH falling from 124 +/- 49.2 to 29.3 +/- 15.4 mU/L; P < 0.01), the THF+allo-THF/THE ratio rose from 0.53 +/- 0.06 to 0.63 +/- 0.07 (P < 0.05). Patients from either group who failed to demonstrate a change in GH levels showed no change in the THF+allo-THF/THE ratio. In vitro studies conducted on cells stably transfected with either the human 11betaHSD1 or 11betaHSD2 complementary DNA and primary cultures of human omental adipose stromal cells expressing only the 11betaHSD1 isozyme indicated a dose-dependent inhibition of 11betaHSD1 oxo-reductase activity with IGF-I, but not GH. Neither IGF-I nor GH had any effect on 11betaHSD2 activity. GH, through an IGF-I-mediated effect, inhibits 11betaHSD1 activity. This reduction in E to F conversion will increase the MCR of F, and care should be taken to monitor the adequacy of function of the hypothalamo-pituitary-adrenal axis in acromegalic subjects and in GH-deficient, hypopituitary patients commencing replacement GH therapy. Conversely, enhanced E to F conversion occurs with a reduction in GH levels; in liver and adipose tissue this would result in increased hepatic glucose output and visceral adiposity, suggesting that part of the phenotype currently attributable to adult GH deficiency may be an indirect consequence of its effect on tissue F metabolism via 11betaHSD1 expression.
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PMID:Modulation of 11beta-hydroxysteroid dehydrogenase isozymes by growth hormone and insulin-like growth factor: in vivo and in vitro studies. 1056 68

The UV-visible and electron paramagnetic resonance (EPR) spectra of MCR(red1), the catalytically active state of methyl-coenzyme M reductase, are almost identical to those observed when free coenzyme F430 or its pentamethyl ester (F430M) are reduced to the Ni(I) valence state. Investigations and proposals concerning the catalytic mechanism of MCR were therefore based on MCR(red1) containing Ni(I)F430 until, in a recent report, Tang et al. (J. Am. Chem. Soc. 2002, 124, 13242) interpreted their resonance Raman data and titration experiments as indicating that, in MCR(red1), coenzyme F430 is not only reduced at the nickel center but at one of the C=N double bonds of the hydrocorphinoid macrocycle as well. To resolve this contradiction, we have investigated the stoichiometry of the reduction of coenzyme F430 pentamethyl ester (F430M) by three independent methods. Spectroelectrochemistry showed clean reduction to a single product that exhibits the UV-vis spectrum typical for MCR(red1). In three bulk electrolysis experiments, 0.96 +/- 0.1 F/mol was required to generate the reduced species. Reduction with decamethylcobaltocene in tetrahydrofuran (THF) consumed 1 mol of (Cp)(2)Co/mol of F430M, and the stoichiometry of the reoxidation of the reduced form with the two-electron oxidant methylene blue was 0.46 +/- 0.05 mol of methylene blue/mol of reduced F430M. These experiments demonstrate that the reduction of coenzyme F430M to the species having almost identical UV-vis and EPR spectra as MCR(red1) is a one-electron process and therefore inconsistent with a reduction of the macrocycle chromophore.
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PMID:Direct determination of the number of electrons needed to reduce coenzyme F430 pentamethyl ester to the Ni(I) species exhibiting the electron paramagnetic resonance and ultraviolet-visible spectra characteristic for the MCR(red1) state of methyl-coenzyme M reductase. 1457 Apr 85

We propose a new DFT-based mechanism for methane production using the full F430 cofactor of MCR (methyl-coenzyme M reductase) along with a coordinated O=CH2CH2C(H)NH2C(H)O (surrogate for glutamine) as a model of the active site for conversion of CH3SCoM(-) (CH3SCH2CH2SO3(-)) + HSCoB to methane plus the corresponding heterodisulfide. The cycle begins with the protonation of F430, either on Ni or on the C-ring nitrogen of the tetrapyrrole ring, both of which are nearly equally favorable. The C-ring protonated form is predicted to oxidatively add CH3SCoM(-) to give a 4-coordinate Ni center where the Ni moves out of the plane of the four ring nitrogens. The movement of Ni (and the attached CH3 and SCH2CH2SO3(2-) ligands) toward the SCoB(-) (deprotonated HSCoB) cofactor allows a 2c-3e interaction to form between the two sulfur atoms. The release of the heterodisulfide yields a Ni(III) center with a methyl group attached. The concerted elimination of methane, where the methyl group coordinated to Ni abstracts the proton from the C-ring nitrogen, has a very small calculated activation barrier (5.4 kcal/mol). The NPA charge on Ni for the various reaction steps indicates that the oxidation state changes occur largely on the attached ligands.
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PMID:A new mechanism for methane production from methyl-coenzyme M reductase as derived from density functional calculations. 1824 3

This study was conducted to investigate the possible relationship between a molecular biological investigation and water quality parameters in monitoring groundwater pollution at the immediate boundary of uncontrolled landfills and their downgradient aquifers, which may consequently facilitate unbiased monitoring for the sites. Two closed landfills, Jicksan and Taejang in Korea, were chosen for this study, where the diversity of the microbial community was characterized and three specific genes, i.e. nirS (nitrite reductase coding gene), cnorB (nitric oxide reductase coding gene) and MCR (methyl coenzyme M reductase coding gene), were quantified. The quantified genes were then compared with conventional water quality parameters. From the analyzed DNA sequences, Proteobacteria phylum was most dominantly observed. A quantitative analysis revealed that the copy numbers (gene abundance) of denitrification enzyme coding genes, i.e. nirS gene and cnorB gene in Jicksan (J) site, are seven and four times, respectively, higher than Taejang (T) site. This simply implied that denitrification was possibly higher in J site than T site. In addition, a methane production enzyme coding gene, i.e. MCR, in a J1 bore immediately bordering the sources in the J site showed the greatest concentration, but it was precipitously decreased in the downgradient direction toward the outer boundary of landfill. A comparative investigation between the copy numbers of three genes, i.e. nirS, cnorB, and MCR, and conventional monitoring parameters, i.e. Cl-, alkalinity, TOC, NH3-N, and NO2-N, showed that they had overall correlation as given by more than 0.99 of the squared correlation coefficient (R2) for almost all of the concerned bores. It was concluded that the comparison between the molecular biological investigation and the conventional groundwater monitoring parameters showed good relationship between them, so that both tools could be more efficiently used for assessing the levels of contamination and prediction of the fate of pollutants, rather than being applied separately.
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PMID:Comparision of nirS, cnorB, MCR genes against water quality parameters to monitor uncontrolled landfills. 1869 20

The nature of the nickel center in the ox1 form of methylcoenzyme M reductase (MCR(ox1)), the enzyme that catalyzes the last step of biological methanogenesis, has long been controversial. A recent pulse electron paramagnetic resonance (EPR) study suggested a Ni(III)-thiolate, or equivalently a high-spin Ni(II) thiyl radical, description. The MCR(ox1) hyperfine parameters are best interpreted in terms of a Ni d(x(2)-y(2) ) SOMO, although a pure d(x(2)-y(2) ) SOMO does not explain the fact that about 7 % of the spin-density resides on the sulfur. The key goals of this DFT (PW91) study were to judge whether the Ni(III)-thiolate description is chemically sensible and, if so, to provide a detailed molecular orbital (MO) description of MCR(ox1). An Ni(III)-thiolate description was indeed found to be reasonable and was obtained as the ground state for symmetrized (C(s)) oxaporphyrin-, pyriporphyrin-, and isoporphyrin-based model complexes, as well as for a more realistic, biomimetic model. The model calculations yielded a number of insights, key among which are the following: 1) Although the SOMO topology may be viewed as d(x(2)-y(2) )-like, this MO also has a substantial amount of metal d(z(2) ) character, allowing it to overlap with a thiolate sigma lone pair, which would otherwise be orthogonal. 2) In one case (isoporphyrin), we were able to exploit the symmetry of the molecule to independently optimize the (d(x(2)-y(2) ))(1) and (d(z(2) ))(1) Ni(III) states, which turned out to be very close in energy. 3) The near-degeneracy of these two states provides an elegant explanation for the tendency of these two orbitals to hybridize. Admixture of substantial d(z(2) ) character into the d(x(2)-y(2) )-type SOMO of our most realistic model of MCR(ox1) results in a small but distinct spin population of about 0.04 on the sulfur, apparently nicely confirming the conclusions derived from the pulse EPR study. Other pure functionals also confirm this picture, although the hybrid functional B3LYP yields a spin-density profile that is clearly at odds with the EPR study.
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PMID:Models of the ox1 state of methylcoenzyme M reductase: where are the electrons? 1881 56

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