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:1.12.7.2 (hydrogenase)
3,522 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Different species of methanogenic bacteria growing on CO(2) and H(2) were shown to remove CO added to the gas phase. Rates up to 0.2 mumol of CO depleted/min per 10 ml of culture containing approximately 7 mg of cells (wet weight) were observed. Methanobacterium thermoautotrophicum was selected for further study based on its ability to grow rapidly on a completely mineral medium. This species used CO as the sole energy source by disproportionating CO to CO(2) and CH(4) according to the following equation: 4CO + 2H(2)O --> 1CH(4) + 3CO(2). However, growth was slight, and the growth rate on CO was only 1% of that observed on H(2)/CO(2). Growth only occurred with CO concentrations in the gas phase of lower than 50%. Growth on CO agrees with the finding that cell-free extracts of M. thermoautotrophicum contained both an active factor 420 (F(420))-dependent hydrogenase (7.7 mumol/min per mg of protein at 35 degrees C) and a CO-dehydrogenating enzyme (0.2 mumol/min per mg of protein at 35 degrees C) that catalyzed the reduction of F(420) with CO. The properties of the CO-dehydrogenating enzyme are described. In addition to F(420), viologen dyes were effective electron acceptors for the enzyme. The apparent K(m) for CO was higher than 1 mM. The reaction rate increased with increasing pH and displayed an inflection point at pH 6.7. The temperature dependence of the reaction rate followed the Arrhenius equation with an activation energy (DeltaHdouble dagger) of 14.1 kcal/mol (59.0 kJ/mol). The CO dehydrogenase activity was reversibly inactivated by low concentrations of cyanide (2 muM) and was very sensitive to inactivation by oxygen. Carbon monoxide dehydrogenase of M. thermoautotrophicum exhibited several characteristic properties found for the enzyme of Clostridium pasteurianum but differed mainly in that the clostridial enzyme did not utilize F(420) as the electron acceptor.
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PMID:Carbon monoxide oxidation by methanogenic bacteria. 2 Nov 59

Extracts of aerobically, CO-autotrophically grown cells of Pseudomonas carboxydovorans were shown to catalyze the oxidation of CO to CO(2) in the presence of methylene blue, pyocyanine, thionine, phenazine methosulfate, or toluylene blue under strictly anaerobic conditions. Viologen dyes and NAD(P)(+) were ineffective as electron acceptors. The same extracts catalyzed the oxidation of formate and of hydrogen gas; the spectrum of electron acceptors was identical for the three substrates, CO, formate, and H(2). The CO- and the formate-oxidizing activities were found to be soluble enzymes, whereas hydrogenase was membrane bound exclusively. The rates of oxidation of CO, formate, and H(2) were measured spectrophotometrically following the reduction of methylene blue. The rate of carbon monoxide oxidation followed simple Michaelis-Menten kinetics; the apparent K(m) for CO was 45 muM. The reaction rate was maximal at pH 7.0, and the temperature dependence followed the Arrhenius equation with an activation energy (DeltaH(0)) of 35.9 kJ/mol (8.6 kcal/mol). Neither free formate nor hydrogen gas is an intermediate of the CO oxidation reaction. This conclusion is based on the differential sensitivity of the activities of formate dehydrogenase, hydrogenase, and CO dehydrogenase to heat, hypophosphite, chlorate, cyanide, azide, and fluoride as well as on the failure to trap free formate or hydrogen gas in coupled optical assays. These results support the following equation for CO oxidation in P. carboxydovorans: CO + H(2)O --> CO(2) + 2 H(+) + 2e(-) The CO-oxidizing activity of P. carboxydovorans differed from that of Clostridium pasteurianum by not reducing viologen dyes and by a pH optimum curve that did not show an inflection point.
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PMID:Oxidation of carbon monoxide in cell extracts of Pseudomonas carboxydovorans. 3 64

The gene encoding a protein containing a putative [6Fe-6S] prismane cluster has been cloned from Desulfovibrio vulgaris (Hildenborough) and sequenced. The gene encodes a polypeptide composed of 553 amino acids (60,161 Da). The DNA-derived amino acid sequence was partly confirmed by N-terminal sequencing of the purified protein and of fragments of the protein generated by CNBr cleavage. Furthermore, the C-terminal sequence was verified by digestion with carboxypeptidases A and B. The polypeptide contains nine Cys residues. Four of these residues are gathered in a Cys-Xaa2-Cys-Xaa7-Cys-Xaa5-Cys motif located towards the N-terminus of the protein. No relevant sequence similarity was found with other proteins, including those with high-spin Fe-S clusters (nitrogenase, hydrogenase), with one significant exception: the stretch containing the first four Cys residues spans two submotifs, Cys-Xaa2-Cys and Lys-Gly-Xaa-Cys-Gly, separated by 11 residues, that are also present in high-spin Fe-S cluster containing CO dehydrogenase. Western-blot analysis demonstrates cross-reactivity of antibodies raised against the purified protein both in Desulfovibrio strains and other sulfate-reducing bacteria. Hybridization of the cloned gene with genomic DNA of several other Desulfovibrio species indicates that homologous sequences are generally present in the genus Desulfovibrio.
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PMID:The primary structure of a protein containing a putative [6Fe-6S] prismane cluster from Desulfovibrio vulgaris (Hildenborough). 133 51

The reduction potentials of two relatively high-molecular-mass enzymes were determined directly at an edge pyrolytic graphite electrode by using square-wave voltammetry. The equilibrium reduction potential versus standard hydrogen electrode was determined for Clostridium pasteurianum hydrogenase I (E'0 = -377 +/- 10 mV; molecular mass 60 kDa) and Rhodospirillum rubrum carbon monoxide dehydrogenase (E'0 = -418 +/- 7 mV; molecular mass 62 kDa). The reduction potential of each enzyme was pH-independent, and one electron was transferred per redox centre. The reduction potential was determined to be identical for the CO dehydrogenase, semi-apo-(CO dehydrogenase), and CO dehydrogenase with carbonyl sulphide (COS) or cyanide bound. The electron-transferring efficiency of CO dehydrogenase was affected by two inhibitors, COS and cyanide, as indicated by a diminished analytic current.
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PMID:Direct electrochemical studies of hydrogenase and CO dehydrogenase. 163 98

Strains of Clostridium thermoaceticum were tested for H2- and CO-dependent growth in a defined medium containing metals, minerals, vitamins, cysteine-sulfide, CO2-bicarbonate, and H2 or CO. Ten of the thirteen strains tested grew at the expense of H2 and CO, and C. thermoaceticum ATCC 39073 was chosen for further study. The doubling times for H2- and CO-dependent growth under chemolithotrophic conditions (the defined medium with nicotinic acid as sole essential vitamin and sulfide as sole reducer) were 25 and 10 h, respectively. Product stiochiometries for chemolithotrophic cultures approximated: 4.1H2 + 2.4CO2----CH3COOH + 0.1 cell C + 0.3 unrecovered C and 6.8CO----CH3COOH + 3.5CO2 + 0.4 cell C + 0.9 unrecovered C. H2-dependent growth produced significantly higher acetate concentrations per unit of biomass synthesized than did CO- or glucose-dependent growth. In contrast, the doubling time for H2-dependent growth under chemolithotrophic conditions (the defined medium without vitamins and sulfide as sole reducer) by Acetogenium kivui ATCC 33488 was 2.7 h; as a sole energy source, CO was not growth supportive for A. kivui. The YH2 values for A. kivui and C. thermoaceticum were 0.91 and 0.46 g of cell dry weight per mol of H2 consumed, respectively; the YCO value for C. thermoaceticum was 1.28 g of cell dry weight per mol of CO consumed. The specific activities of hydrogenase and CO dehydrogenase in both acetogens were influenced by the energy source utilized for growth and were significantly lower in C. thermoaceticum than in A. kivui. With extracts of H2-cultivated cells and benzyl viologen as electron acceptor, the Vmax values for hydrogenase from C. thermoaceticum and A. kivui were 155.7 and 1,670 micromoles of H2 oxidized per min mg of protein, respectively; the Vmax values for CO dehydrogenase from C. thermoaceticum and A. kivui were 90.6 and 2,973 micromoles of CO oxidized per min per mg of protein, respectively.
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PMID:Characterization of the H2- and CO-dependent chemolithotrophic potentials of the acetogens Clostridium thermoaceticum and Acetogenium kivui. 237 65

Exposure of the photosynthetic bacterium Rhodospirillum rubrum to carbon monoxide led to increased carbon monoxide dehydrogenase and hydrogenase activities due to de novo protein synthesis of both enzymes. Two-dimensional gels of [35S]methionine-pulse-labeled cells showed that induction of CO dehydrogenase synthesis was rapidly initiated (less than 5 min upon exposure to CO) and was inhibited by oxygen. Both CO dehydrogenase and the CO-induced hydrogenase were inactivated by oxygen in vivo and in vitro. In contrast to CO dehydrogenase, the CO-induced hydrogenase was 95% inactivated by heating at 70 degrees C for 5 min. Unlike other hydrogenases, this CO-induced hydrogenase was inhibited only 60% by a 100% CO gas phase.
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PMID:Regulation of carbon monoxide dehydrogenase and hydrogenase in Rhodospirillum rubrum: effects of CO and oxygen on synthesis and activity. 249 85

A ferredoxin has been isolated from the thermophilic methanogen Methanococcus thermolithotrophicus. The native protein was a monomer exhibiting a molecular weight of 7,262, calculated from the amino acid composition. Its absorption spectrum had two maxima at 390 and 283 nm, with an absorbance ratio A390/A283 of 0.79. The absorption at 390 nm (E = 29 mM-1 cm-1) and the content of iron of the protein are in agreement with the presence of two 4Fe-4S clusters in M. thermolithotrophicus ferredoxin. Its amino acid composition showed the presence of eight cysteine residues, which is the required number of cysteines for the binding of two 4Fe-4S clusters. The protein was characterized by the lack of histidine, arginine, and leucine and a high content of valine. It was unusually stable to high temperatures but not to oxygen. The ESR spectrum of the protein in the oxidized state showed a minor signal at g = 2.01, corresponding to an oxidized 3Fe-4S cluster. The protein, which was difficult to reduce with dithionite or reduced mediators, exhibited in its reduced state a spectrum typical of two interacting reduced 4Fe-4S clusters. M. thermolithotrophicus ferredoxin functioned as an electron acceptor for the CO dehydrogenase complex with an extract free of ferredoxin. No reaction was detected with F420 or hydrogenase.
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PMID:Isolation, characterization, and biological activity of the Methanococcus thermolithotrophicus ferredoxin. 254 Jan 48

In Pseudomonas carboxydovorans, CO dehydrogenase and hydrogenase were found in association with the cytoplasmic membrane in a weakly bound and a tightly bound pool. The pools could be experimentally distinguished on the basis of resistance to removal by washes in low-ionic-strength buffer. The tightly bound pool of the enzymes could be differentially solubilized under conditions leaving the electron transport system intact and with the nondenaturing zwitterionic detergent 3-(3-cholamidopropyl) dimethylammonio 1-propane-sulfonic acid (CHAPS) and the nonionic detergent dodecyl beta-D-maltoside. In vitro reconstitution of depleted membranes with the corresponding supernatants containing CO dehydrogenase led to binding of the enzyme and to reactivation of respiratory activities with CO. The reconstitution reaction required cations with effectiveness which increased with increasing ionic charge: monovalent (Li+), divalent (Mg2+, Mn2+), or trivalent (Cr3+, La3+). Reconstitution of depleted membranes with CO dehydrogenase was specific for CO-grown bacteria. Cytoplasmic membranes from H2- or heterotrophically grown Pseudomonas carboxydovorans had no affinity for CO dehydrogenase at all, indicating the absence of the physiological electron acceptor of the enzyme, which presumably is cytochrome b561, or another membrane anchor.
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PMID:Removal of CO dehydrogenase from Pseudomonas carboxydovorans cytoplasmic membranes, rebinding of CO dehydrogenase to depleted membranes, and restoration of respiratory activities. 280 5

An inactive, Ni-deficient form of carbon monoxide (CO) dehydrogenase [carbon-monoxide:(acceptor) oxidoreductase; EC 1.2.99.2], designated apo-CO dehydrogenase, accumulated in Rhodospirillum rubrum when cells were grown in the absence of Ni and treated with CO. In vivo, both CO dehydrogenase activity and hydrogenase activity increased several hundred fold upon addition of 2 microM NiCl2. Apo-CO dehydrogenase was purified to homogeneity and differed from holo-CO dehydrogenase only in its activity and Ni content, containing less than 0.2 mol of Ni per mol of protein, and a specific activity of 35 mumol of CO per min per mg. Optimal in vitro activation of purified apo-CO dehydrogenase resulted in an enzyme with a specific activity of 2640 mumol of CO per min per mg. No additional enzymes or low molecular weight cofactors were required for activation. Apo-CO dehydrogenase was not activated by MgCl2, MnCl2, CuCl2, ZnCl2, CoCl2, or Na2MoO4. 63Ni was incorporated into apo-CO dehydrogenase during activation. The electron paramagnetic resonance (EPR) spectra of dithionite-reduced apo- and holo-enzyme were identical, indicating that, in the reduced state, the Fe-S centers observed by EPR are unchanged in the apo-enzyme.
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PMID:Nickel-deficient carbon monoxide dehydrogenase from Rhodospirillum rubrum: in vivo and in vitro activation by exogenous nickel. 282 76

Cell extracts from acetate-grown Methanosarcina thermophila contained CO-oxidizing:H2-evolving activity 16-fold greater than extracts from methanol-grown cells. Following fractionation of cell extracts into soluble and membrane components, CO-dependent H2 evolution and CO-dependent methyl-coenzyme M methylreductase activities were only present in the soluble fraction, but addition of the membrane fraction enhanced both activities. A b-type cytochrome(s), present in the membrane fraction, was linked to a membrane-bound hydrogenase. CO-oxidizing:H2-evolving activity was reconstituted with: (i) CO dehydrogenase complex, (ii) a ferredoxin, and (iii) purified membranes with associated hydrogenase. The ferredoxin was a direct electron acceptor for the CO dehydrogenase complex. The ferredoxin also coupled CO oxidation by CO dehydrogenase complex to metronidazole reduction.
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PMID:Ferredoxin requirement for electron transport from the carbon monoxide dehydrogenase complex to a membrane-bound hydrogenase in acetate-grown Methanosarcina thermophila. 327 28


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