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)

Eight strains of Rhizobium lacking hydrogenase uptake (Hup) activity and 17 transconjugant strains carrying the hup cosmids pHU1, pHU52, or pHU53 (G. R. Lambert, M. A. Cantrell, F. J. Hanus, S. A. Russell, K. R. Haddad, and H. J. Evans, Proc. Natl. Acad. Sci. USA, 82:3232-3236, 1985) were screened for Hup activity and the presence of immunologically detectable hydrogenase polypeptides. Crude extracts of these strains were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot analysis with affinity-purified antibodies against the two subunits of purified hydrogenase (Mr 60,000 and 30,000). Derepressed transconjugants carrying the cosmid pHU52 were Hup+ and contained detectable levels of both hydrogenase subunit polypeptides. Non-derepressed strains, Hup- parent strains, and strains carrying cosmids other than pHU52 did not express Hup activity and contained no immunologically detectable protein. These data provide further evidence for the essential involvement of the smaller (Mr 30,000) subunit in the expression of hydrogenase activity in Rhizobium japonicum and suggest that the determinants for hydrogenase subunit synthesis are present on pHU52.
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PMID:Further evidence that two unique subunits are essential for expression of hydrogenase activity in Rhizobium japonicum. 390 36

Rhizobium japonicum hydrogenase was purified to homogeneity from soybean root nodules by four column chromatography steps after solubilization from membranes by treatment with a nonionic detergent. The specific activity was from 40 to 65 mumol H2 oxidized min-1 mg protein-1 and was increased 450-fold relative to that in bacteroids. The yield of activity was from 7 to 12%. The molecular weight of the native enzyme was 104,000 as determined by sucrose density gradient centrifugation. Electrophoresis in the presence of sodium dodecyl sulfate revealed two subunits with molecular weights of 64,000 and 35,000, indicating an alpha beta subunit structure. The amino acid content of the protein indicated 20 cysteine residues. Analysis of the metal content indicated 0.59 +/- 0.06 mol Ni/mol hydrogenase and 6.5 +/- 1.2 mol Fe/mol hydrogenase. Antisera prepared to the hydrogenase cross-reacted with the enzyme in bacteroid extracts at all stages of the purification but did not cross-react with extracts of Alcaligenes eutrophus grown under chemolithotrophic conditions. The similarity of rhizobial hydrogenase to the particulate hydrogenases of A. eutrophus and A. latus is discussed.
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PMID:Rhizobium japonicum hydrogenase: purification to homogeneity from soybean nodules, and molecular characterization. 391 48

Acryloyl-CoA reductase, a presumably previously unknown soluble enzyme, is present in Clostridium kluyveri. It catalyses the reduction of the carbon-carbon double bond of acryloyl-CoA or ethyl vinyl ketone and other alpha, beta-unsaturated carbonyl compounds at the expense of reduced methylviologen. On the basis of a Vmax/Km ratio, which is at least 18 times higher than that for the next best substrate (E)-2-butenoyl-CoA, the enzyme is called acryloyl-CoA reductase. A purity of over 90% was achieved. The apparent molecular mass, as determined by gel chromatography, is 28.4 kDa. Dodecyl sulfate gel electrophoresis shows subunits with a molecular mass of 14.2 kDa. Based on a molecular mass of 28.4 kDa about 1.5 mol FMN have been observed. Less than 0.2 g-atom iron per mol protein were determined. Ferredoxin or flavodoxin seem to be able to carry electrons from hydrogenase to the acryloyl-CoA reductase. The addition of hydrogen to the alpha-carbon of ethyl vinyl ketone occurs from the re-side.
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PMID:Purification and some properties of an acryloyl-CoA reductase of Clostridium kluyveri. 406 66

Several dissimilatory, sulfate-reducing bacteria were isolated from the rumen fluid of sheep fed purified diets containing sulfate. One isolate, strain D, was selected for characterization. This organism is a nonsporeforming, obligately anaerobic, mesophilic, nonmotile, gram-negative, straight rod. Cell-free extracts show absorption maxima for cytochrome c(3) and desulfoviridin, characteristic of Desulfovibrio. Carbohydrates, as a sole carbon source, will support growth. Lactate supports growth in the presence of sulfate, not in its absence, whereas glucose or pyruvate support growth either in the presence or absence of sulfate. The isolate has a deoxyribonucleic acid base composition of 61.2% guanine plus cytosine, which is similar to that of several other species of Desulfovibrio; however, it differs from previously described species in morphology, motility, and carbon source utilization. Cell-free extracts of this bacterium exhibit adenosine 5'-triphosphate-sulfurylase, adenosine-5'-phosphosulfate-reductase, and hydrogenase activity. After incubation of cell-free extracts with adenine 5'-triphosphate and (35)SO(4) (2-), adenosine-5'-phosphosulfate rather than 3'-phosphoadenosine-5'-phosphosulfate was shown to be labeled, indicating that the pathway of sulfate reduction in this organism is similar to that of other dissimilatory sulfate reducers. This is the first report of a Desulfovibrio sp. isolated from the rumen.
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PMID:Sulfate reduction by a Desulfovibrio species isolated from sheep rumen. 447 25

A type of iron-bound protein was isolated from Clostridium botulinum by a modification of the method used for isolating ferredoxin from C. pasteurianum. This method involved acetone and diethylaminoethyl cellulose treatments followed by ammonium sulfate fractionation. The protein exhibited maximal absorption in the ultraviolet region near 260 mmu. Portions of the isolated iron protein were separated by disc electrophoresis and, following specific iron-bound protein staining, showed a positive reaction in the same position on the gel column as was first demonstrated by use of cell-free extract. Evidence accumulated by use of a cell-free extract of C. botulinum suggests that pyruvate is metabolized through a phosphoroclastic system as demonstrated in other clostridia. It is probable that ferredoxin is an electron mediator between pyruvic oxidase and hydrogenase for hydrogen evolution and acetyl phosphate formation.
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PMID:Isolation and role of nonheme iron protein in Clostridium botulinum. 486 57

From salt flats on the Galapagos Islands, two strains of a red photosynthetic bacterium were isolated and identified as Ectothiorhodospira mobilis, an organism first described by Pelsh in 1937. The cells are curved in a short spiral, 0.7 to 1.0 mu wide and 2.0 to 4.8 mu long. They are motile by a polar tuft of flagella. Cells contain several large stacks of lamellar membranes, carrying the pigments bacteriochlorophyll a and carotenoids of the spirillo xanthin series. Cell division occurs by binary fission, not budding. The organism is strictly anaerobic and obligately photosynthetic. Its ability to grow well with sulfide, sulfur, thiosulfate, or sulfite as photosynthetic H donors puts it taxonomically in the Thiorhodaceae. During growth with sulfide, elementary sulfur is deposited outside the cells in the medium and disappears during further growth. A limited number of organic carbon compounds can be utilized as hydrogen donors in place of inorganic sulfur compounds. Under these conditions, sulfate can serve as the sulfur source. The enzymes catalase and hydrogenase are present. The newly isolated strains require vitamin B(12). They also require a salinity of 2 to 3% NaCl, but they are not extreme halophiles. The organism is not identical with any of the species listed in Bergey's Manual.
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PMID:Ectothiorhodospira mobilis Pelsh, a photosynthetic sulfur bacterium depositing sulfur outside the cells. 565 91

A soluble hydrogenase from the methanogenic bacterium, Methanosarcina barkeri (DSM 800) has been purified to apparent electrophoretic homogeneity, with an overall 550-fold purification, a 45% yield and a final specific activity of 270 mumol H2 evolved min-1 (mg protein)-1. The hydrogenase has a high molecular mass of approximately equal to 800 kDa and subunits with molecular masses of approximately equal to 60 kDa. The enzyme is stable to heating at 65 degrees C and to exposure to air at 4 degrees C in the oxidized state for periods up to a week. The overall stability of this enzyme is compared with other hydrogenase isolated from strict anaerobic sulfate-reducing bacteria. Ms. barkeri hydrogenase shows an absorption spectrum typical of a non-heme iron protein with maxima at 275 nm, 380 nm and 405 nm. A flavin component, identified as FMN or riboflavin was extracted under acidic conditions and quantified to approximately one flavin molecule per subunit. In addition to this component, 8-10 iron atoms and 0.6-0.8 nickel atom were also detected per subunit. The electron paramagnetic resonance (EPR) spectrum of the native enzyme shows a rhombic signal with g values at 2.24, 2.20 and approximately equal to 2.0. probably due to nickel which is optimally measured at 40 K but still detectable at 77 K. In the reduced state, using dithionite or molecular hydrogen as reductants, at least two types of g = 1.94 EPR signals, due to iron-sulfur centers, could be detected and differentiated on the basis of power and temperature dependence. Center I has g values at 2.04, 1.90 and 1.86, while center II has g values at 2.08, 1.93 and 1.85. When the hydrogenase is reduced by hydrogen or dithionite the rhombic EPR species disappears and is replaced by other EPR-active species with g values at 2.33, 2.23, 2.12, 2.09, 2.04 and 2.00. These complex signals may represent different nickel species and are only observable at temperatures higher than 20 K. In the native preparation, at high temperatures (T greater than 35 K) or in partially reduced samples, a free radical due to the flavin moiety is observed. The EPR spectrum of reduced hydrogenase in 80% Me2SO presents an axial type of spectrum only detectable below 30 K.
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PMID:Purification, characterization and redox properties of hydrogenase from Methanosarcina barkeri (DSM 800). 608 41

A c3 type cytochrome has been purified from the thermophilic, non-spore-forming, sulfate-reducing bacterium Thermodesulfobacterium commune. The purified protein was homogeneous as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, gel filtration, and isoelectric focusing. A pI of 6.83 was observed. The molecular weight of the cytochrome was estimated to be ca. 13,000 from both gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The hemoprotein exhibited absorption maxima at 530, 408.5, and 351 nm in the oxidized form and 551.5 (alpha band), 522.5 (beta band), and 418.5 nm (gamma band) in the reduced form. The extinction coefficients of T. commune cytochrome c3 were 130,000, 74,120, and 975,000 M-1 cm-1 at 551.5, 522.5, and 418.5 nm, respectively. It contains four hemes per molecule, on the basis of both the iron estimation and the extinction coefficient value of its pyridine hemochrome. The amino acid composition showed the presence of eight cysteine residues involved in heme binding. T. commune cytochrome c3 had low threonine, serine, and glycine contents and high glutamic acid and hydrophobic residue contents. The electrochemical study of T. commune cytochrome c3 by cyclic voltammetry and differential pulse polarography has shown that the cytochrome system behaves like a reversible system. Four redox potential values at Eh1 = -0.140 +/- 0.010 V, Eh2 = Eh3 = Eh4 = -0.280 +/- 0.010 V have been determined. T. commune cytochrome c3, which acts as the physiological electron carrier of hydrogenase, is similar in most respects to the multiheme low-potential cytochrome c3 which is characteristic of the genus Desulfovibrio.
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PMID:Characterization of cytochrome c3 from the thermophilic sulfate reducer Thermodesulfobacterium commune. 609 Mar 84

A hydrogenase from a new species of sulfate reducing bacterium has been isolated and characterized. In contrast to other hydrogenases isolated from Desulfovibrio, this enzyme is found in the cytoplasmic fraction rather than in the periplasm. The specific activity of the enzyme, as measured in the hydrogen evolution assay, is twice as high as the specific activity of the hydrogenase from D. gigas. It also differentiates itself from the periplasmic Desulfovibrio hydrogenases by being more active in the hydrogen evolution rather than in the hydrogen uptake assay. The enzyme was shown to contain 0.9 atoms of nickel, 11 atoms of iron and 10 atoms of labile sulfide per mole of enzyme. It exhibits an unusually low intensity of the g = 2.31 nickel EPR signal in the isolated enzyme but shows a normal intensity for the g = 2.19 nickel EPR signal when reduced under hydrogen.
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PMID:A cytoplasmic nickel-iron hydrogenase with high specific activity from Desulfovibrio multispirans sp. N., a new species of sulfate reducing bacterium. 609 50

The reduction of CO2 or any other methanogenic substrate to methane serves the same function as the reduction of oxygen, nitrate or sulfate to more reduced products. These exergonic reactions are coupled to the production of usable energy generated through a charge separation and a protonmotive-force-driven ATPase. For the understanding of how methanogens derive energy from C-1 unit reduction one must study the biochemistry of the chemical reactions involved and how these are coupled to the production of a charge separation and subsequent electron transport phosphorylation. Data on methanogenesis by a variety of organisms indicates ubiquitous use of CH3-S-CoM as the final electron acceptor in the production of methane through the methyl CoM reductase and of 5-deazaflavin as a primary source of reducing equivalents. Three known enzymes serve as catalysts in the production of reduced 5-deazaflavin: hydrogenase, formate dehydrogenase and CO dehydrogenase. All three are potential candidates for proton pumps. In the organisms that must oxidize some of their substrate to obtain electrons for the reduction of another portion of the substrate to methane (e.g., those using formate, methanol or acetate), the latter two enzymes may operate in the oxidizing direction. CO2 is the most frequent substrate for methanogenesis but is the only substrate that obligately requires the presence of H2 and hydrogenase. Growth on methanol requires a B12-containing methanol-CoM methyl transferase and does not necessarily need any other methanogenic enzymes besides the methyl-CoM reductase system when hydrogenase is present. When bacteria grow on methanol alone it is not yet clear if they get their reducing equivalents from a reversal of methanogenic enzymes, thus oxidizing methyl groups to CO2. An alternative (since these and acetate-catabolizing methanogens possess cytochrome b) is electron transport and possible proton pumping via a cytochrome-containing electron transport chain. Several of the actual components of the methanogenic pathway from CO2 have been characterized. Methanofuran is apparently the first carbon-carrying cofactor in the pathway, forming carboxy-methanofuran. Formyl-FAF or formyl-methanopterin (YFC, a very rapidly labelled compound during 14C pulse labeling) has been implicated as an obligate intermediate in methanogenesis, since methanopterin or FAF is an essential component of the carbon dioxide reducing factor in dialyzed extract methanogenesis. FAF also carries the carbon at the methylene and methyl oxidation levels.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:The bioenergetics of methanogenesis. 623 47


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