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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)

Formaldehyde hydrogenase and formate dehydrogenase were purified 130-fold and 19-fold respectively from Candida boidinii grown on methanol. The final enzyme preparations were homogenous as judged by acrylamide gel electrophoresis and by sedimentation in an ultracentrifuge. The molecular weights of the enzymes were determined by sedimentation equilibrium studies and calculated as 80000 and 74000 respectively. Dissociation into subunits was observed by treatment with sodium dodecylsulfate. The molecular weights of the polypeptide chains were estimated to be 40000 and 36000 respectively. The NAD-linked formaldehyde dehydrogenase specifically requires reduced glutathione for activity. Besides formaldehyde only methylglyoxal served as a substrate but no other aldehyde tested. The Km values were found to be 0.25 mM for formaldehyde, 1.2 mM for methylglyoxal, 0.09 mM for NAD and 0.13 mM for glutathione. Evidence is presented which demonstrates that the reaction product of the formaldehyde-dehydrogenase-catalyzed oxidation of formaldehyde is S-formylglutathione rather than formate. The NAD-linked formate dehydrogenase catalyzes specifically the oxidation of formate to carbon dioxide. The Km values were found to be 13 mM for formate and 0.09 mM for NAD.
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PMID:Purification and properties of formaldehyde dehydrogenase and formate dehydrogenase from Candida boidinii. 124 77

S organism, an anaerobic gram-negative rod, which is one of two bacterial species isolated from the culture known as "Methanobacillus omelianskii," ferments ethanol to acetate and H(2). The present study shows that extracts of this organism contain ferredoxin and produce acetate from acetaldehyde via aldehyde: ferredoxin oxidoreductase activity. Electrons generated in the reaction are given off as H(2) by a previously demonstrated ferredoxin-linked hydrogenase system. Extracts were shown to contain good phosphotransacetylase and acetokinase activities, but no mechanism of adenosine triphosphate generation during acetaldehyde conversion to acetate could be detected. No evidence could be obtained for coenzyme A (CoASH) or phosphate requirement or for formation of acetyl CoA or acetyl phosphate.
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PMID:Ferredoxin-dependent conversion of acetaldehyde to acetate and H 2 in extracts of S organism. 455 84

The pathways of glucose and pyruvate metabolism in Spirochaeta litoralis, a free-living, strictly anaerobic marine spirochete, were studied. Addition of 0.2 to 0.4 M NaCl (final concentration) to suspending buffers prevented cell lysis and was necessary for gas evolution from various substrates by cell suspensions. The organism fermented glucose mainly to ethanol, acetate, CO(2), and H(2). Determination of radioactivity in products formed from (14)C-labeled glucose and assays of enzymatic activities in cell extracts indicated that S. litoralis catabolized glucose via the Embden-Meyerhof pathway. A clostridial-type clastic reaction was utilized by the spirochete to degrade pyruvate to acetyl-coenzyme A, CO(2), and H(2). Formation of acetate from acetyl-coenzyme A was catalyzed by phosphotransacetylase and acetate kinase. Nicotinamide adenine dinucleotide-dependent acetaldehyde and alcohol dehydrogenases converted acetyl-coenzyme A to ethanol. A reversible hydrogenase activity was detected in cell extracts. S. litoralis cell extracts contained a rubredoxin similar in spectral properties to other bacterial rubredoxins.
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PMID:Glucose and pyruvate metabolism of Spirochaeta litoralis, an anaerobic marine spirochete. 474 35

Tritrichomonas foetus mutants resistant to metronidazole lack the hydrogenosomal enzymes pyruvate: ferredoxin oxidoreductase and hydrogenase. Hydrogenosomes of these organisms did not oxidize pyruvate or produce ATP in its presence. Elimination of hydrogenosomal metabolism of pyruvate was compensated by an increased rate of glycolysis. The resistant mutants excreted no organic acids and H2 as metabolic end products. Glycolysis of the resistant T. foetus KV1-1MR-100 can be summarized as 1 mol glucose----2 mol ethanol + 2 mol CO2. The parent strain KV1, excreting H2, CO2 and acidic end products, converted about 10% of glucose to ethanol. Both strains produced ethanol from pyruvate through the action of two cytoplasmic enzymes: pyruvate decarboxylase and alcohol dehydrogenase. The specific activity of the former enzyme, catalyzing nonoxidative decarboxylation of pyruvate to acetaldehyde, was nearly seven times higher in the resistant than in the parent strain. Alcohol dehydrogenase reducing acetaldehyde to ethanol was specific to NADPH; it catalyzed the reverse reaction only slowly, and displayed similar activities in both resistant and sensitive trichomonads. Development of anaerobic metronidazole resistance in T. foetus depended on the loss of pyruvate:ferredoxin oxidoreductase as well as on the ability to increase alcoholic fermentation.
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PMID:Metabolic differences between metronidazole resistant and susceptible strains of Tritrichomonas foetus. 637 46

Acetaldehyde was shown to be an irreversible inhibitor of nitrogenase, hydrogenase, CO2 fixation and growth in the cyanobacterium Anabaena cylindrica, but had no effect on photosynthetic electron flow as measured by Methyl Viologen-dependent O2 uptake. The concentration-dependence of the inhibition of nitrogenase and hydrogenase activities was determined, and it was shown that acetaldehyde inhibition poses problems for anaerobic experiments in which the activities of these enzymes are measured in the presence of the frequently used glucose/glucose oxidase/catalase/ethanol O2 trap. It is suggested that acetaldehyde may find use as an inhibitor in experiments designed to separate electron flow through the photosystems from consequent fixation of CO2 and N2.
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PMID:The effects of acetaldehyde on nitrogenase, hydrogenase and photosynthesis in the cyanobacterium Anabaena cylindrica. 641 Oct 74

The in vitro and in vivo effects of selected pharmacologic agents on cytosolic aldehyde dehydrogenase in the adult rat testicles were studied. Naltrexone, a narcotic antagonist, significantly inhibited epididymal but not testicular aldehyde dehydrogenase in vitro. Greater in vitro inhibition of aldehyde dehydrogenase in the epididymal preparation was determined in the presence of diphenhydramine, a histamine H1-receptor antagonist and by amantadine hydrochloride, an antiviral/antiparkinsonian agent. The latter drug also inhibited testicular aldehyde dehydrogenase from corresponding controls. The in vitro inhibition was noncompetitive as evaluated by reciprocal plots of the velocity of the reaction as a function of substrate concentrations. Short-term oral administration of a biologically active metabolite of flutamide, a nonsteroidal antiandrogen, significantly induced testicular aldehyde dehydrogenase without concomitant changes in epididymal aldehyde dehydrogenase from respective controls. Conversely, administration of ethinylestradiol, an estrogenic steroid, inhibited testicular but not epididymal aldehyde hydrogenase in the rat.
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PMID:Effect of pharmacologic interventions on aldehyde dehydrogenase in the rat testicles. 718 91

The strictly anaerobic archaeon Thermococcus strain ES-1 was recently isolated from near a deep-sea hydrothermal vent. It grows at temperatures up to 91 degrees C by the fermentation of peptides and reduces elemental sulfur (S(o)) to H2S. It is shown here that the growth rates and cell yields of strain ES-1 are dependent upon the concentration of S(o) in the medium, and no growth was observed in the absence of S(o). The activities of various catabolic enzymes in cells grown under conditions of sufficient and limiting S(o) concentrations were investigated. These enzymes included alcohol dehydrogenase (ADH); formate benzyl viologen oxidoreductase; hydrogenase; glutamate dehydrogenase; alanine dehydrogenase; aldehyde ferredoxin (Fd) oxidoreductase; formaldehyde Fd oxidoreductase; and coenzyme A-dependent, Fd-linked oxidoreductases specific for pyruvate, indolepyruvate, 2-ketoglutarate, and 2-ketoisovalerate. Of these, changes were observed only with ADH, formate benzyl viologen oxidoreductase, and hydrogenase, the specific activities of which all dramatically increased in cells grown under S(o) limitation. This was accompanied by increased amounts of H2 and alcohol (ethanol and butanol) from cultures grown with limiting S(o). Such cells were used to purify ADH to electrophoretic homogeneity. ADH is a homotetramer with a subunit M(r) of 46,000 and contains 1 g-atom of Fe per subunit, which, as determined by electron paramagnetic resonance analyses, is present as a mixture of ferrous and ferric forms. No other metals or acid-labile sulfide was detected by colorimetric and elemental analyses. ADH utilized NADP(H) as a cofactor and preferentially catalyzed aldehyde reduction. It is proposed that, under So limitation, ADH reduces to alcohols the aldehydes that are generated by fermentation, thereby serving to dispose of excess reductant.
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PMID:Effects of elemental sulfur on the metabolism of the deep-sea hyperthermophilic archaeon Thermococcus strain ES-1: characterization of a sulfur-regulated, non-heme iron alcohol dehydrogenase. 764 2

The main function of the hydrogenosome, a typical organelle of trichomonads, is to convert malate or pyruvate to H(2), CO(2) and acetate by a pathway associated with ATP synthesis. This pathway relies on activity of iron-sulfur proteins such as pyruvate:ferredoxin oxidoreductase (PFOR), hydrogenase and ferredoxin. To examine the effect of iron availability on proper hydrogenosomal function, the metabolic activity of the hydrogenosome and expression of hydrogenosomal enzymes were compared in Tritrichomonas foetus maintained under iron-rich (150 microM iron nitrilotriacetate) or iron-restricted (180 microM 2,2-dipyridyl) conditions in vitro. The activities of PFOR and hydrogenase, and also production of acetate and H(2), were markedly decreased or absent in iron-restricted trichomonads. Moreover, a decrease in activity of the hydrogenosomal malic enzyme, which is a non-Fe-S protein, was also observed. Impaired function of hydrogenosomes under iron-restricted conditions was compensated for by activation of the cytosolic pathway, mediating conversion of pyruvate to ethanol via acetaldehyde. This metabolic switch was fully reversible. Production of hydrogen by iron-restricted trichomonads was restored to the level of organisms grown under iron-rich conditions within 3 h after addition of 150 microM iron nitrilotriacetate. Protein analysis of purified hydrogenosomes from iron-restricted cells showed decreased levels of proteins corresponding to PFOR, malic enzyme and ferredoxin. Accordingly, these cells displayed decreased steady-state level and synthesis of mRNAs encoding PFOR and hydrogenosomal malic enzyme. These data demonstrate that iron is essential for function of the hydrogenosome, show its involvement in the expression of hydrogenosomal proteins and indicate the presence of iron-dependent control of gene transcription in Tt. foetus.
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PMID:Iron-induced changes in pyruvate metabolism of Tritrichomonas foetus and involvement of iron in expression of hydrogenosomal proteins. 1116 Aug

A series of azadithiolate (adt)-bridged Fe-only hydrogenase model complexes, Fe2(CO)6(mu-adt)C6H4I-4 (1), Fe2(CO)6(mu-adt)C6H4C[triple bond]CR [R = C6H4NO2-4 (2), C6H4CHO-4 (3), C6H4NH2-4 (4), C6H4COOH-4 (5), C6H4COOCH2CH3-4 (6), C6H4F-4 (7), C6H5 (8), C6H4OCH3-4 (9), C6H4N(CH3)2-4 (10)], [Fe2(CO)5(PPh3)(mu-adt)C6H4I-4 (11), and Fe2(CO)5(PPh3)(mu-adt)C6H4C[triple bond]CC6H4NO2-4 (12), have been synthesized in high yields under mild conditions. The linear geometry and rigidity of a triple bond act as an effective bridge to anchor a functionality ranging from electron-donating to electron-accepting, even coordinative groups in the adt model complexes. X-ray crystal analysis of 2, 3, and 6-12 reveals that the model complexes retain the butterfly structure of Fe2S2 model analogues. A rigid phenylacetylene offers excellent control over the distance between the functional group and the active site of Fe2S2 model complexes. The unusual Fe-Fe distance and the angles found in the molecular packing of 6 are originated from the intriguing intermolecular C-H...O and C-H...S interactions. More importantly, electrochemical studies reveal that all of the complexes can catalyze electrochemical reduction of protons to molecular hydrogen, but the reduction potential for the electron-transfer step can be remarkably altered by the functionality R. The electroreductively active nitro group in 2 and 12 displays the enhanced current at a potential substantially less negative than the reduction of [Fe(I)Fe(I)] + e(-) --> [Fe(I)Fe(0)], which is most accessible and becomes the initial step. For complex 3, the second reduction peak for the electron-transfer step involves the contribution from the aldehyde functionality. As the electroreductively inactive groups are incorporated, the reduction process of [Fe(I)Fe(I)] + e(-) --> [Fe(I)Fe(0)] appears first and the second reduction peak for the electron-transfer step from the [Fe(I)Fe(0)] + e(-) --> [Fe(0)Fe(0)] process for 4-10 is clearly observed. Therefore, the order of electron and proton uptake is closely related to the electroreductively active functionality, R. Varying the nature of the functionality R leads to the electron-transfer step changes from the reduction of the electroreductively active R group to the active site of Fe2S2 model complexes subsequently. Accordingly, notwithstanding, acetic acid is too weak to protonate the series of 2-12, different reduction pathways can be followed, and the electrochemically catalyzed behavior may occur at different reduction levels.
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PMID:Facile synthesis and functionality-dependent electrochemistry of Fe-only hydrogenase mimics. 1871 Feb 14

The production of the chemical solvents acetone and butanol by the bacterium Clostridium acetobutylicum was one of the first large-scale industrial processes to be developed, and in the first part of the last century ranked second in importance only to ethanol production. After a steep decline in its industrial use, there has been a recent resurgence of interest in the acetone-butanol-ethanol (ABE) fermentation process, with a particular emphasis on butanol production. In order to generate strains suitable for efficient use on an industrial scale, metabolic engineering is required to alter the AB ratio in favour of butanol, and eradicate the production of unwanted products of fermentation. Using ClosTron technology, a large-scale targeted mutagenesis in C. acetobutylicum ATCC 824 was carried out, generating a set of 10 mutants, defective in alcohol/aldehyde dehydrogenases 1 and 2 (adhE1, adhE2), butanol dehydrogenases A and B (bdhA, bdhB), phosphotransbutyrylase (ptb), acetate kinase (ack), acetoacetate decarboxylase (adc), CoA transferase (ctfA/ctfB), and a previously uncharacterised putative alcohol dehydrogenase (CAP0059). However, inactivation of the main hydrogenase (hydA) and thiolase (thl) could not be achieved. Constructing such a series of mutants is paramount for the acquisition of information on the mechanism of solvent production in this organism, and the subsequent development of industrial solvent producing strains. Unexpectedly, bdhA and bdhB mutants did not affect solvent production, whereas inactivation of the previously uncharacterised gene CAP0059 resulted in increased acetone, butanol, and ethanol formation. Other mutants showed predicted phenotypes, including a lack of acetone formation (adc, ctfA, and ctfB mutants), an inability to take up acids (ctfA and ctfB mutants), and a much reduced acetate formation (ack mutant). The adhE1 mutant in particular produced very little solvents, demonstrating that this gene was indeed the main contributor to ethanol and butanol formation under the standard batch culture conditions employed in this study. All phenotypic changes observed could be reversed by genetic complementation, with exception of those seen for the ptb mutant. This mutant produced around 100 mM ethanol, no acetone and very little (7 mM) butanol. The genome of the ptb mutant was therefore re-sequenced, together with its parent strain (ATCC 824 wild type), and shown to possess a frameshift mutation in the thl gene, which perfectly explained the observed phenotype. This finding reinforces the need for mutant complementation and Southern Blot analysis (to confirm single ClosTron insertions), which should be obligatory in all further ClosTron applications.
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PMID:Targeted mutagenesis of the Clostridium acetobutylicum acetone-butanol-ethanol fermentation pathway. 2298 1


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