A ferredoxin-Iinked sulfite reductase from Clostridium pasteurianum

1971 ◽  
Vol 17 (7) ◽  
pp. 889-895 ◽  
Author(s):  
E. J. Laishley ◽  
Po-Min Lin ◽  
H. D. Peck Jr.
Keyword(s):  
Phosphate Buffer ◽  
Molecular Hydrogen ◽  
Methyl Viologen ◽  
Hydrogen Uptake ◽  
Electron Donors ◽  
Sulfite Reductase ◽  
Clostridium Pasteurianum ◽  
Ph Optimum ◽  
Benzyl Viologen ◽  
Dissimilatory Sulfite Reductase

A soluble sulfite reductase (EC. 1.8.1.2) system is present in cell-free extracts of Clostridium pasteurianum that reduces sulfite to sulfide in the presence of molecular hydrogen. The natural electron donor for this reductase has been found to be ferredoxin, which can be completely replaced by methyl viologen or partially by benzyl viologen. The physiological electron donors NAD, NADP, FMN, and FAD were not active in the transfer of electrons from hydrogen for sulfite reduction. The pH optimum of the sulfite reductase was found to be 7.0 using phosphate buffer, and the extracts reduced nitrite and hydroxyl-amine in addition to sulfite. The stoichiometry of the reaction in terms of hydrogen uptake to sulfide formation was 2.7, which is very close to the theoretical ratio of 3. The above properties of the system indicate that it is an assimilatory rather than a dissimilatory sulfite reductase, as outlined in the Discussion.

The interaction of polymeric viologens with hydrogenases from Desulfovibrio desulfuricans and Clostridium pasteurianum

1979 ◽  
Vol 57 (8) ◽  
pp. 1093-1098 ◽  
Author(s):  
Bernard R. Glick ◽  
William G. Martin ◽  
J. Jean Giroux ◽  
Ross E. Williams
Keyword(s):  
Hydrogen Production ◽  
Methyl Viologen ◽  
Desulfovibrio Desulfuricans ◽  
Electron Donors ◽  
Clostridium Pasteurianum ◽  
Maximal Rate ◽  
Free System ◽  
Electrophoretic Band ◽  
Cell Free System ◽  
A Cell

The interaction between hydrogenases from either Desulfovibrio desulfuricans or Clostridium pasteurianum and electron donors methyl viologen or polymeric viologens was examined. Extracts from each organism contained a single gel electrophoretic band of active hydrogenase. The hydrogenase of D. desulfuricans was much more stable than that of Cl. pasteurianum. With methyl viologen apparent Km and Vm values were 0.5 mM and 0.62 μmol H2/min per milligram protein for the Cl. pasteurianum and 0.7 and 6.2 μmol H2/min per milligram protein, respectively, for the D. desulfuricans enzyme. The hydrogenases bound the polymeric viologens more tightly than methyl viologen, more so for the enzyme of D. desulfuricans than for Cl. pasteurianum. Maximal rate of hydrogen production was less with the polymeric than with methyl viologen. The results suggest that the D. desulfuricans enzyme in conjunction with a polymeric viologen may perform better in a cell-free system aimed at hydrogen production than that from Cl. pasteurianum.

Stable isotope fractionation by Clostridium pasteurianum. 1.34S/32S: inverse isotope effects during SO42− and SO32− reduction

1975 ◽  
Vol 21 (3) ◽  
pp. 235-244 ◽  
Author(s):  
R. G. L. McCready ◽  
E. J. Laishley ◽  
H. R. Krouse
Keyword(s):  
Isotope Fractionation ◽  
Isotope Effects ◽  
Sulfite Reductase ◽  
Clostridium Pasteurianum ◽  
Stages Of Growth ◽  
Dissimilatory Sulfite Reductase ◽  
Stable Isotope Fractionation ◽  
High Concentrations ◽  
Late Stages ◽  
Isotopic Effects

During growth on minimal salts – sucrose media supplemented with various concentrations (10−4–10−2 M) of sodium sulfate, Clostridium pasteurianum grew at a normal rate and only evolved sulfide in late stages of growth on 10−2 M SO42−. The evolved sulfide was slightly enriched in 34S as compared to the medium sulfur. On the other hand, sulfide was evolved during growth on all concentrations of sulfite tested. Large normal and inverse isotopic effects were observed in the evolved sulfide during SO32− reductions. In contrast, the intracellular sulphur showed much smaller fractionations. The complexity of the isotopic patterns suggests that a dissimilatory sulfite reductase system may be induced by high concentrations of sulfite.

scholarly journals Purification of the membrane-bound hydrogenase of Escherichia coli

1979 ◽  
Vol 183 (1) ◽  
pp. 11-22 ◽  
Author(s):  
M W W Adams ◽  
D O Hall
Keyword(s):  
Escherichia Coli ◽  
Methyl Viologen ◽  
Hydrophobic Interaction Chromatography ◽  
H2 Evolution ◽  
Ph Optimum ◽  
Benzyl Viologen ◽  
Electron Carrier ◽  
Membrane Bound ◽  
High Concentrations ◽  
Electron Carriers

The membrane-bound hydrogenase (EC class 1.12) of aerobically grown Escherichia coli cells was solubilized by treatment with deoxycholate and pancreatin. The enzyme was further purified to electrophoretic homogeneity by chromoatographic methods, including hydrophobic-interaction chromatography, with a yield of 10% as judged by activity and an overall purification of 2140-fold. The hydrogenase was a dimer of identical subunits with a mol.wt. of 113,000 and contained 12 iron and 12 acid-labile sulphur atoms per molecule. The epsilon 400 was 49,000M-1 . cm-1. The hydrogenase catalysed both H2 evolution and H2 uptake with a variety of artificial electron carriers, but would not interact with flavodoxin, ferredoxin or nicotinamide and flavin nucleotides. We were unable to identify any physiological electron carrier for the hydrogenase. With Methyl Viologen as the electron carrier, the pH optimum for H2 evolution and H2 uptake was 6.5 and 8.5 respectively. The enzyme was stable for long periods at neutral pH, low temperatures and under anaerobic conditions. The half-life of the hydrogenase under air at room temperature was about 12 h, but it could be stabilized by Methyl Viologen and Benzyl Viologen, both of which are electron carriers for the enzyme, and by bovine serum albumin. The hydrogenase was strongly inhibited by carbon monoxide (Ki = 1870Pa), heavy-metal salts and high concentrations of buffers, but was resistant to inhibition by thiol-blocking and metal-complexing reagents. These aerobically grown E. coli cells lacked formate hydrogenlyase activity and cytochrome c552.

Pyridine–Adenine Dinucleotide Transhydrogenase Activity in Cells Cultured from Rat Hepatoma

1972 ◽  
Vol 50 (5) ◽  
pp. 447-456 ◽  
Author(s):  
C. De Luca ◽  
R. P. Gioeli
Keyword(s):  
Phosphate Buffer ◽  
Pyridine Nucleotide ◽  
Maximum Activity ◽  
Ph Optimum ◽  
Apparent Lack ◽  
Minimal Deviation ◽  
Ph Range ◽  
Pyridine Nucleotide Transhydrogenase ◽  
Rat Hepatoma ◽  
Cells Cultured

Preparations from cells cultured from a minimal-deviation hepatoma in the rat exhibit pyridine nucleotide transhydrogenase (NAD(P)H: NAD(P) oxidoreductase, EC 1.6.1.1) activity. The pH optimum, its release by digitonin, and its apparent lack of dependence on steroids for activity tentatively classify it as a transhydrogenase of the type first described for animal tissue.Enzyme preparations from digitonin-treated homogenates were very unstable. The time necessary for the loss of one-half the activity was 16–18 h when the enzyme was stored at 5 °C; this was reduced to 4 h when storage was in polycarbonate tubes.The enzyme apparently transferred hydrogen directly and with equal ease from NADH to both the 3-acetyl-pyridine and thionicotinamide analogues of NAD. Half-saturation values for NAD and its acetylpyridine analogue were 0.99 × 10−5 M and 3.55 × 10−4 M, respectively. The enzyme exhibited its maximum activity in phosphate buffer at pH 5.8. It was inhibited by 50–60% over the pH range 7.0–8.5 in Tris buffer. This could be reversed by dithiothreitol; reversal was complete between pH 8.0 and 8.5.

scholarly journals The biosynthesis of valine from isobutyrate by Peptostreptococcus elsdenii and Bacteroides ruminicola

1971 ◽  
Vol 121 (3) ◽  
pp. 431-437 ◽  
Author(s):  
Milton J. Allison ◽  
J. L. Peel
Keyword(s):  
Molecular Hydrogen ◽  
Photosynthetic Bacteria ◽  
Deae Cellulose ◽  
Cell Protein ◽  
Acetyl Phosphate ◽  
Acid Fraction ◽  
Benzyl Viologen ◽  
Electron Carrier ◽  
Thiamin Pyrophosphate ◽  
Reductive Carboxylation

1. Growing cultures of Peptostreptococcus elsdenii and Bacteroides ruminicola incorporate 14C from [1-14C]isobutyrate into the valine of cell protein. With P. elsdenii some of the 14C is also incorporated into leucine. 2. Crude cell-free extracts of both organisms in the presence of glutamine, carbon dioxide and suitable sources of energy and electrons incorporate 14C from [1-14C]isobutyrate into valine but not into leucine. 3. With extracts of P. elsdenii treated with DEAE-cellulose the reaction is dependent on ATP, CoA, thiamin pyrophosphate, molecular hydrogen and a low-potential electron carrier (ferredoxin, flavodoxin or benzyl viologen). 4. The same extracts incorporate 14C from NaH14CO3 into valine in the presence of isobutyrate plus ATP, CoA, glutamine and ferredoxin; isobutyryl-CoA or isobutyryl phosphate plus CoA will replace the isobutyrate plus CoA and ATP. With acetyl phosphate in place of isobutyryl phosphate, 14C is incorporated into alanine. With isovalerate or 2-methylbutyrate in place of isobutyrate, 14C is incorporated into leucine and isoleucine respectively. 5. When carrier 2-oxoisovalerate is added to the carboxylating system 14C from [1-14C]isobutyrate passes into the oxo acid fraction. 6. It is concluded that these two organisms form valine from isobutyrate by the sequence isobutyrate→isobutyryl-CoA→2-oxoisovalerate→valine and that the reductive carboxylation of isobutyrate is catalysed by a system similar to the pyruvate synthetase of clostridia and photosynthetic bacteria.

Pyruvate:ferredoxin oxidoreductase in Campylobacter species

1995 ◽  
Vol 41 (2) ◽  
pp. 198-201 ◽  
Author(s):  
James A. Daucher ◽  
Noel R. Krieg
Keyword(s):  
Clostridium Pasteurianum ◽  
Benzyl Viologen ◽  
Minimal Inhibitory Concentrations ◽  
Ferredoxin Oxidoreductase ◽  
Crude Extracts ◽  
Reducing Activity ◽  
Pyruvate Ferredoxin Oxidoreductase ◽  
Pyruvate:Ferredoxin Oxidoreductase ◽  
Campylobacter Species

Twelve strains representing 11 Campylobacter species were tested by two methods to see whether pyruvate:ferredoxin oxidoreductase, which occurs mainly in anaerobes, was also present in these oxygen-respiring microaerophiles. Crude extracts exhibited a pyruvate-dependent reduction of benzyl viologen and, to a lesser extent, metronidazole. Addition of Clostridium pasteurianum ferredoxin enhanced the metronidazole-reducing activity. All strains were inhibited by metronidazole, with minimal inhibitory concentrations ranging from 3 to 25 μg/mL. The results suggest that pyruvate: ferredoxin oxidoreductase is characteristic of the genus.Key words: Campylobacter, microaerophilic, metronidazole, pyruvate.

scholarly journals Evaluation of the Bioelectrochemical Approach and Different Electron Donors for Biological Trichloroethylene Reductive Dichlorination

Toxics ◽  
2022 ◽  
Vol 10 (1) ◽  
pp. 37
Author(s):  
Edoardo Dell’Armi ◽  
Marta Maria Rossi ◽  
Lucia Taverna ◽  
Marco Petrangeli Papini ◽  
Marco Zeppilli
Keyword(s):  
Reductive Dechlorination ◽  
Molecular Hydrogen ◽  
Batch Reactor ◽  
Electron Donors ◽  
Utilization Efficiency ◽  
Batch Reactors ◽  
Organic Substrates ◽  
Chlorinated Aliphatic Hydrocarbons ◽  
Low Solubility ◽  
Dechlorination Reaction

Trichloroethylene (TCE) and more in general chlorinated aliphatic hydrocarbons (CAHs) can be removed from a contaminated matrix thanks to microorganisms able to perform the reductive dechlorination reaction (RD). Due to the lack of electron donors in the contaminated matrix, CAHs’ reductive dechlorination can be stimulated by fermentable organic substrates, which slowly release molecular hydrogen through their fermentation. In this paper, three different electron donors constituted by lactate, hydrogen, and a biocathode of a bioelectrochemical cell have been studied in TCE dechlorination batch experiments. The batch reactors evaluated in terms of reductive dechlorination rate and utilization efficiency of the electron donor reported that the bio-electrochemical system (BES) showed a lower RD rate with respect of lactate reactor (51 ± 9 µeq/d compared to 98 ± 4 µeq/d), while the direct utilization of molecular hydrogen gave a significantly lower RD rate (19 ± 8 µeq/d), due to hydrogen low solubility in liquid media. The study also gives a comparative evaluation of the different electron donors showing the capability of the bioelectrochemical system to reach comparable efficiencies with a fermentable substrate without the use of other chemicals, 10.7 ± 3.3% for BES with respect of 3.5 ± 0.2% for the lactate-fed batch reactor. This study shows the BES capability of being an alternative at classic remediation approaches.

Studies on the Mechanism of NAD-photoreduction by Chromatophores of the Facultative Phototroph, Rhodopseudomonas capsulata

1969 ◽  
Vol 24 (1) ◽  
pp. 67-76 ◽  
Author(s):  
J.-H. Klemme
Keyword(s):  
Nicotinamide Adenine Dinucleotide ◽  
Molecular Hydrogen ◽  
Reaction System ◽  
Electron Donors ◽  
Rhodopseudomonas Capsulata ◽  
Antimycin A ◽  
Inhibitory Effects ◽  
Carbonyl Cyanide ◽  
Cyclic Photophosphorylation ◽  
Energy Dependent

The light-driven and the ATP-driven reduction of nicotinamide adenine dinucleotide (NAD) catalyzed by the chromatophore fraction of Rhodopseudomonas capsulata was investigated. Efficient electron donors for the photoreduction of NAD are molecular hydrogen and succinate. In the ATP-dependent reaction system, succinate is a more efficient electron donor than H2. The energydependent NAD-reduction is driven by ATP, but not by pyrophosphate or ADP. Oligomycin stimulates the NAD-photoreductions and completely inhibits the ATP-driven NAD-reductions. Rotenone and piericidin A are inhibitors for both the light-driven and the ATP-driven NAD-reductions. Antimycin A is an inhibitor only for the light-driven reductions. The H2-linked NAD-photoreduction is less sensitive to these inhibitors and to the uncoupler desaspidin than the succinate-linked reduction. Atebrine, carbonyl cyanide-m-chlorophenylhydrazone, 2,4-dinitrophenol and phenazonium methosulfate are inhibitors for the light-driven and the ATP-driven reductions. Some of the compounds used as inhibitors of the NAD-reduction were also investigated with concerns to their inhibitory effects on cyclic photophosphorylation and O2-linked oxidations of reduced NAD, succinate and H2. Based on the results of these inhibitor studies, the relationships between cyclic photophosphorylation, light-induced noncyclic electron transport and energy-dependent NAD-reduction are discussed.

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