Inhibitory effect of molecular hydrogen on C2H2-reducing activity in Anabaena 7120 preilluminated in red or blue light

Molecular hydrogen inhibits nitrogenase activity in Anabaena pre-illuminated with red or blue light. The inhibitory effect of molecular hydrogen decreased in the presence of oxygen and several electron acceptors. When NH₄Cl and urea were added simultaneously with molecular hydrogen, marked synergistic inhibitory effects took place. The inhibitory effect of molecular hydrogen disappeared or was weakened after the suppression of hydrogenase activity. The addition of O₂ and electron acceptors to systems showed no enhancing effect on the C₂H₂-reducing activity.     

Structure-function relationship in hemoproteins: The role of cytochrome c 3 in the reduction of colloidal sulfur by sulfate-reducing bacteria

Cytochromes c ₃ of different strains of sulfatereducing bacteria have been purified and tested for their capacity to reduce colloidal sulfur to hydrogen sulfide. The results are in good agreement with the activities reported for the whole cells. Cytochrome c ₃ is the sulfur reductase of some strains of sulfate-reducing bacteria such as Desulfovibrio desulfuricans Norway 4 and sulfate-reducing bacterium strain 9974 from which the sulfur reductase activity can be purified with the cytochrome c ₃. In contrast, Desulfovibrio vulgaris Hildenborough cytochrome c ₃ is inhibited by the product of the reaction namely hydrogen sulfide. Chloramphenicol has no effect on the sulfur reductase activity of D. desulfuricans Norway 4 when resting cells grown on lactate-sulfate medium are put in the presence of colloidal sulfur. This shows that the sulfur reductase activity is constitutive and corresponds to the fact that colloidal sulfur grown cells do not contain more cytochrome c ₃ (or another sulfur reductase) than lactate-sulfate-grown cells.     

Catalytic De/Hydrogenation in Mg by Co‐Doped Ni and VOx on Active Carbon: Extremely Fast Kinetics at Low Temperatures and High Hydrogen Capacity

A multi‐component catalyst Ni‐VO_x/AC (VO_x is comprised of V₂O₅ and VO₂, x = 2.18) was synthesized by a wet impregnation method. The synthesized Ni‐VO_x/AC shows a superior catalytic effect on de/hydrogenation of Mg. The MgH₂+Ni‐VO_x/AC composites can absorb 6.2 wt.‐% hydrogen within only 1 min at 150 °C under a hydrogen pressure of 2 MPa and desorb 6.5 wt.‐% hydrogen within 10 min at 300 °C under an initial hydrogen pressure of 1 KPa, which overcomes a critical barrier for practical use of Mg as a hydrogen storage material. A significant decrease of activation energy (E_a) indicates that Ni‐VO_x/AC catalyst is highly efficient for Mg de/hydrogenation, which may be ascribed to the synergistic effect of bimetals (metal oxides) and nanocarbon.     

The structure of triple-stranded G-2C polynucleotide helices.

The thermal stability of a new polynucleotide complex has been used to establish the hydrogen-bonding structure of three-stranded C-G·CH⁺ helices. In the Hoogsteen structure, the 8NH₂ group of 8NH₂GMP can form a third hydrogen bond to the CH⁺ strand, but in the alternative structure, the 8NH₂ group can form no interbase hydrogen bonds. For the new complex, 8NH₂GMP·2 poly(C), a transition temperature of 80°C is observed under conditions in which the corresponding complex formed with 5′-GMP has a T_m of 20°C. We conclude from this 60° elevation of transition temperature that a third hydrogen bond is formed by the 8NH₂ group and that the structure must have Hoogsteen bonding. In order to be compatible with this structure in regular helices formed by U,C copolymers, A·2U bonding would also have to have a Hoogsteen structure.     

Induction of H2-Uptake and Nitrogenase Activities in the Cyanobacterium Anabaena variabilis ATCC 29413: Effects of Hydrogen and Organic Substrate

A comparative study of the development of uptake hydrogenase and nitrogenase activities in cells of the cyanobacterium Anabaena variabilis was performed. The induction of heterocysts is followed by the induction of both in vivo hydrogen uptake and nitrogenase activities. Interestingly, a low but significant H₂-uptake [2–7 μmoles of H₂ · mg⁻¹ (Chl a) · h⁻¹] occurs in cultures with no heterocysts and with no nitrogenase activity. A slight stimulatory effect (30–40%) of H₂ on in vivo H₂-uptake was observed during the early stages of nitrogenase induction. However, exogenous H₂ does not further stimulate the induction of in vivo hydrogen uptake observed during heterocyst differentiation. Similarly, organic carbon (fructose) did not influence the induction of either in vivo hydrogen uptake or nitrogenase activities. Exogenous fructose supports higher in vivo hydrogen uptake and nitrogenase activities when the cells enter late exponential phase of growth. Received: 22 November 1995 / Accepted: 22 December 1995     

The use of 3-amino-1,2,4-triazole to investigate the short-term effects of oxygen toxicity on carbon assimilation by Pisum sativum seedlings

To study the in vivo short-term effect of hydrogen peroxide on plant metabolism, 2 mol m^?3 3-amino-1,2,4-triazole, a catalase inhibitor, was applied through the transpiration stream to Pisum sativum seedlings, and gas exchange characteristics, ascorbate peroxidase, glutathione reductase and catalase activities, and levels of hydrogen peroxide and formate were determined. Carbon dioxide assimilation rates were inhibited after the addition of aminotriazole: photorespiratory conditions exacerbated this inhibition. Carbon dioxide response curves showed that aminotriazole reduced both the RuBP regeneration rate and the efficiency of the carboxylation reaction of Rubisco. Catalase activity was completely inhibited 200 min after the application of this inhibitor, but no concomitant increase in H₂O₂ concentration was found. Under enhanced photorespiratory conditions, H₂O₂ concentrations increased. This suggests that under normal environmental conditions hydrogen peroxide is metabolized via alternative mechanisms. The aminotriazole treatment had no effect on the ascotbate peroxidase and glutathione reductase activities, but caused a substantial increase in the formate pool size. These results suggest that hydrogen peroxide is metabolized by reacting with glyoxylate to produce formate and CO₂. The increased production of formate may reduce the flow of carbon through the normal photorespiratory pathway and may also be used anaplerotically as a precursor of products of 1-C metabolism other than serine. This would prevent the return of photorespiratory carbon to the RPP pathway, leading to a smaller RuBP pool size which would in turn result in a decrease in carboxylation conductance (carboxylation efficiency) and regeneration rate of RuBP.     

Effect of hydrogen peroxide on antioxidant enzyme activities in Saccharomyces cerevisiae is strain-specific

The effect of hydrogen peroxide on the survival and activity of antioxidant and associated enzymes in Saccharomyces cerevisiae has been studied. A difference found in the response of wild-type yeast strains treated with hydrogen peroxide was probably related to the different protective effects of antioxidant enzymes in these strains. Exposure of wild-type YPH250 cells to 0.25 mM H₂O₂ for 30 min increased activities of catalase and superoxide dismutase (SOD) by 3.4-and 2-fold, respectively. However, no activation of catalase in the EG103 strain, as well as of SOD in the YPH98 and EG103 wild strains was detected, which was in parallel to lower survival of these strains under oxidative stress. There is a strong positive correlation (R ² = 0.95) between activities of catalase and SOD in YPH250 cells treated with different concentrations of hydrogen peroxide. It is conceivable that catalase would protect SOD against inactivation caused by oxidative stress and vice versa. Finally, yeast cell treatment with hydrogen peroxide can lead to either a H₂O₂-induced increase in activities of antioxidant and associated enzymes or their decrease depending on the H₂O₂ concentration used or the yeast strain specificity. Published in Russion in Biokhimiya, 2006, Vol. 71, No. 9, pp. 1243–1252.     

Cyanide assimilation in Rhizobium ORS 571: influence of the nitrogenase catalyzed hydrogen production on the efficiency of growth

When cyanide is gradually added to a nitrogenfixing culture, Rhizobium ORS 571 is capable of assimilating large amounts of cyanide using its nitrogenase. Under these conditions the molar growth yield on succinate (Y _succ) increases from 27 at the start of cyanide addition to 38 at the end. The respiratory chain of cells grown at a concentration of 7 mM cyanide is still very sensitive to cyanide. The increase in growth yield is explained by a decrease in hydrogen production by nitrogenase as soon as cyanide is assimilated. This is confirmed by calculating the influence of hydrogen production on Y _succ. Hydrogen production by nitrogenase has a greater influence on growth yields than the presence or absence of hydrogenase activity. At the end of cyanide addition when all cell nitrogen is synthesized from cyanide and no nitrogen fixation occurs, nitrogenase will be in a very oxidized state.     

Solid-state conformations of α-Aminoisobutyryl-L-prolyl sequences: Crystal structure of Boc-Aib-L-Pro-OBzl

The protected dipeptide Boc-Aib-Pro-OBzl, C₂₁H₃₀N₂O₅, crystallizes in the orthorhombic space group P2₁2₁2₁, with a = 12.820, b = 10.529, c = 16.548Å, and Z = 4. The crystal structure has been solved by direct methods and refined to an R value of 0.074 for 1352 reflections. The Boc-Aib-Pro-OBzl molecule has been shown to adopt an unfolded conformation in the solid state with ?_Aib = 50.5°, Ψ_Aib = 45.3°, ?_Pro = ?64.6°, and Ψ_Pro = 148.1°. The result is in marked contrast with the reported crystal structure of Cbz-Aib-Pro-NHMe, which adopts an intramolecularly hydrogen-bonded β-turn conformation. Comparison with 13 reported conformations of Aib-Pro sequences in the crystalline state revealed that the Aib-Pro sequence adopts an unfolded conformation if the residue that immediately follows the dipeptide sequence possesses no hydrogen available for hydrogen bonding, while a β-turn conformation is preferred if the Pro residue is followed by an NH group. Correlation between pyrrolidine ring puckering of the Pro residue and main-chain conformation in Aib-Pro sequences is discussed.     

Possible accumulation of non-active molecules of catalase and superoxide dismutase in S. cerevisiae cells under hydrogen peroxide induced stress

The effect of hydrogen peroxide on the activities of catalase and superoxide dismutase (SOD) in S. cerevisiae has been studied under different experimental conditions: various H₂O₂ concentrations, time exposures, yeast cell densities and media for stress induction. The yeast treatment with 0.25–0.50 mM H₂O₂ led to an increase in catalase activity by 2–3-fold. At the same time, hydrogen peroxide caused an elevation by 1.6-fold or no increase in SOD activity dependently on conditions used. This effect was cancelled by cycloheximide, an inhibitor of protein synthesis in eukaryotes. Weak elevation of catalase and SOD activities in cells treated with 0.25–0.50 mM H₂O₂ found in this study does not correspond to high level of synthesis of the respective enzyme molecules observed earlier by others. It is well known that exposure of microorganisms to low sublethal concentrations of hydrogen peroxide leads to the acquisition of cellular resistance to a subsequent lethal oxidative stress. Hence, it makes possible to suggest that S. cerevisiae cells treated with low sublethal doses of hydrogen peroxide accumulate non-active stress-protectant molecules of catalase and SOD to survive further lethal oxidant concentrations.     

The utilization of molecular hydrogen by the blue-green alga Anabaena cylindrica

The blue-green alga Anabaena cylindrica is found to consume molecular hydrogen in a hydrogenase dependent reaction. This hydrogen uptake proceeds in the dark and is strictly dependent on oxygen, thus representing a Knallgas reactions. Its rate is almost as high as that of the endogenous respiration in Anabaena. Studies with inhibitors reveal that hydrogen is utilized via the complete respiratory chain providing additional energy for the alga. CO plus C₂H₂ completely block the Knallgas reaction which explains the previously reported considerable increase in the total H₂ formation representing the difference between the nitrogenase-dependent H₂-evolution and the reutilization of the gas catalysed by the hydrogenase in intact Anabaena.H₂ is able to support the C₂H₂-reduction in the dark in a reaction again strictly dependent on oxygen. Moreover, H₂ is also consumed in experiments carried out under far red light and in the presence of dichlorophenyl-dimenthyl-urea (DCMU) where the energy for nitrogen fixation is no longer provided by respiration but by cyclic photophosphorylation. Under these conditions, H₂ is found to supply electrons for the formation of C₂H₄ from C₂H₂ in a reaction no longer dependent on the presence of oxygen. Moreover, in these experiments, the presence of H₂ stabilizes the C₂H₂-reduction activity against the deleterious effect of oxygen.Thus, this communication provides evidence for a triplicate function of the H₂-uptake catalysed by hydrogenase in intact Anabaena which is (a) to provide energy by the Knallgas reaction, (b) to supply reducing equivalents for nitrogenase, (c) to protect nitrogenase from damage by oxygen.Abbreviations DCMU N-(3,4-dichlorophenyl)N,N-dimethylurea - DNP 2-4-dinitrophenol - FCCP carbonylcyanid-p-trifluormethoxyphenyl-hydrazone(=p-CF₃-CCP) - Chl chlorophyll     

Decolorization of melanin by lignin peroxidase from<Emphasis Type="Italic">Phanerochaete chrysosporium</Emphasis>

Melanin was decolorized by lignin peroxidase fromPhanerochaete chrysosporium. This decolorization reaction showed a Michaelis-Mentens type relationship between the decolorization rate and concentration of two substrates: melanin and hydrogen peroxide. Kinetic constants of the decolorization reaction were 0.1 OD₄₇₅/min (V _max) and 99.7 mg/L (K _m) for melanin and 0.08 OD₄₇₅/min (V _max) and 504.9 μM (K _m) for hydrogen peroxide, respectively. Depletion of hydrogen peroxide interrupted the decolorization reaction, indicating the essential requirement of hydrogen peroxide. Pulsewise feeding of hydrogen peroxide continued the decolorizing reaction catalyzed by lignin peroxidase. These results indicate that enzymatic decolorization of melanin has applications in the development of new cosmetic whitening agents.     

Biohydrogen production from wheat straw hydrolysate by dark fermentation using extreme thermophilic mixed culture

Hydrolysate was tested as substrate for hydrogen production by extreme thermophilic mixed culture (70°C) in both batch and continuously fed reactors. Hydrogen was produced at hydrolysate concentrations up to 25% (v/v), while no hydrogen was produced at hydrolysate concentration of 30% (v/v), indicating that hydrolysate at high concentrations was inhibiting the hydrogen fermentation process. In addition, the lag phase for hydrogen production was strongly influenced by the hydrolysate concentration, and was prolonged from approximately 11 h at the hydrolysate concentrations below 20% (v/v) to 38 h at the hydrolysate concentration of 25% (v/v). The maximum hydrogen yield as determined in batch assays was 318.4 ± 5.2 mL‐H₂/g‐sugars (14.2 ± 0.2 mmol‐H₂/g‐sugars) at the hydrolysate concentration of 5% (v/v). Continuously fed, and the continuously stirred tank reactor (CSTR), operating at 3 day hydraulic retention time (HRT) and fed with 20% (v/v) hydrolysate could successfully produce hydrogen. The hydrogen yield and production rate were 178.0 ± 10.1 mL‐H₂/g‐sugars (7.9 ± 0.4 mmol H₂/g‐sugars) and 184.0 ± 10.7 mL‐H₂/day L_reactor (8.2 ± 0.5 mmol‐H₂/day L_reactor), respectively, corresponding to 12% of the chemical oxygen demand (COD) from sugars. Additionally, it was found that toxic compounds, furfural and hydroxymethylfurfural (HMF), contained in the hydrolysate were effectively degraded in the CSTR, and their concentrations were reduced from 50 and 28 mg/L, respectively, to undetectable concentrations in the effluent. Phylogenetic analysis of the mixed culture revealed that members involved hydrogen producers in both batch and CSTR reactors were phylogenetically related to the Caldanaerobacter subteraneus, Thermoanaerobacter subteraneus, and Thermoanaerobacterium thermosaccharolyticum. Biotechnol. Bioeng. 2010;105: 899–908. © 2009 Wiley Periodicals, Inc.     

Structurally symmetrical,easily polarizable hydrogen bonds between side chains in proteins and proton-conducting mechanisms. III

(L -Cys)_n, (L -Lys)_n, and (L -Glu)_n were studied by ir spectroscopy in terms of their degree of deprotonation or protonation. It is shown that structurally symmetrical, easily polarizable SH ?S^? ? ^?S ?HS, N⁺H ?N ? N ?H⁺N, and OH ?O^? ? ^?O ?HO hydrogen bonds are formed between the side chains. The different wave number distributions of the ir continua caused by these hydrogen bonds show that the barrier in the double-minimum proton potential decreases in the series of these hydrogen bonds. The stability of these hydrogen bonds against hydration increases in this series. The OH ?O^? ? ^?O ?HO bonds are not broken by small amounts of water. With (L -Cys)_n the formation of easily polarizable hydrogen bonds and a β-structure–coil transition are strongly interdependent. As a result of this coupling effect, the β-structure–coil transition becomes cooperative. With (L -Glu)_n, the formation of the polarizable hydrogen bonds and the observed conformational change are independent processes. The (L -Glu)_n conformation changes from α-helix to coil only if more than 80% of the residues are deprotonated. Finally, on the basis of the various types of easily polarizable hydrogen bonds, charge shifts in active centers of enzymes and the proton-conducting mechanism through hydrophobic regions of biological membranes are discussed.     

Proton transfer and polarizability of hydrogen bonds formed between cysteine and lysine residues

(L -Cys)_n + N-base systems and (L -Cys)_n + (L -Lys)_n systems were studied by ir spectroscopy. It is shown that in the water-free systems, SH ?N ? S^? ?H⁺N hydrogen bonds are formed. With the (L -Cys)_n + N-base systems, both proton-limiting structures in the SH ?N ? S^? ?H⁺N bonds have equal weight when the pK_a of the protonated N-base is 2 pK_a units larger than that of (L -Cys)_n. The same is true with the water-free (L -Cys)_n + (L -Lys)_n system. Thus, with regard to the type of proton potentials present, these hydrogen bonds are proton-transfer hydrogen bonds showing very large proton polarizabilities. This is confirmed by the occurrence of continua in the ir spectra. Small amounts of water open these hydrogen bonds and increase the transfer of the proton to (L -Lys)_n. In the (L -Lys)_n + N-base systems, with increasing proton transfer the backbone of (L -Cys)_n changes from antiparallel β-structure to coil. In (L -Cys)_n + (L -Lys)_n, the conformation is determined by the (L -Lys)_n conformation and changes depending on the chain length of (L -Lys)_n. Finally, the reactivity increase in the active center of fatty acid synthetase, which should be caused by the shift of a proton, is discussed on the basis of the great proton polarizability of the cysteine–lysine hydrogen bonds.     

Studies on the partially uncoupled oxidation of tetrahydropterins by phenylalanine hydroxylase

The uncoupled portion of the partially uncoupled oxidation of tetrahydropterins by phenylalanine hydroxylase can be described by the same model as we have recently derived for the fully uncoupled reaction (Davis, M.D. and Kaufman, S. (1989) J. Biol. Chem.264, 8585–8596). Although essentially no hydrogen peroxide is formed during the fully coupled oxidation of tetrahydrobiopterin or 6-methyltetrahydropterin by phenylalanine hydroxylase when phenylalanine is the amino acid substrate, significant amounts of hydrogen peroxide are formed during the partially uncoupled oxidation of 6-methyltetrahydropterin whenpara-fluorophenylalanine orpara-chlorophenylalanine are used in place of phenylalanine. Similarly, during the partially uncoupled oxidation of the unsubstituted pterin, tetrahydropterin, even in the presence of phenylalanine, hydrogen peroxide formation is detected. The 4a-carbinolamine tetrahydropterin intermediate has been observed during the fully uncoupled tyrosine-dependent oxidations of tetrahydropterin and 6-methyltetrahydropterin by lysolecithin-activated phenylalanine hydroxylase, suggesting that this species is also a common intermediate for uncoupled oxidations by this enzyme.Abbreviations BH₄ 6-[dihydroxypropyl-(L-erythro)-5,6,7,8-tetrahydropterin (tetrahydrobiopterin) - 6MPH₄ 6-methyl-5,6,7,8-tetrahydropterin - PH₄ 5,6,7,8-tetrahydropterin - BH₃OH 4a-hydroxytetrahydropterin (4a-carbinolamine) - qBH₂ quinonoid dihydrobiopterin - q6MPH₂ quinonoid dihydro-6-methylpterin - qPH₂ quinoid dihydropterin - PAH phenylalanine hydroxylase - DHPR dihydropteridine reductase - PHS phenylalanine hydroxylase stimulating enzyme which is 4a-carbinolamine dehydratase - SOD superoxide dismutase - HPLC high performance liquid chromatography - R.T. retention time Special issue dedicated to Dr. Santiago Grisolia.     

Relative efficiency of nitrogen fixation and the occurrence of hydrogenase in pea root nodules

Summary The apparent K_m(hydrogen) for uptake of hydrogen by pea root nodules was determined. This enabled the concentration gradient necessary for the evolution of hydrogen to be calculated for nodules with no hydrogenase activity. This indicated that hydrogen inhibition of nitrogenase is not likely to be the cause of the low relative efficiency of legume root nodules. The factors that affect electron allocation between protons and nitrogen in nitrogenase are reviewed and it is concluded that there must be some as yet unknown factor that affects electron distribution inRhizobium nitrogenase. One possibility is put forward and considered. A strain ofRhizobium was used that was found to possess hydrogenase activity in combination with pea variety Feltham First but not with variety Meteor. The control of this enzyme is briefly discussed.     

Isolation and physiological characterization of Syntrophus buswellii strain GA from a syntrophic benzoate-degrading,strictly anaerobic coculture

A stable, syntrophic benzoate-degrading bacterial consortium was enriched from sewage sludge. It oxidized benzoate or 3-phenylpropionate to acetate, H₂ and CO₂. As hydrogen scavengers Methanospirillum hungatei and Desulfovibrio sp. were present. The benzoate-degrading bacteria of this syntrophic culture and of Syntrophus buswelli were able to grow with benzoate/crotonate or crotonate alone in the absence of a hydrogen-utilizing partner organism. If crotonate was the only substrate, acetate and butyrate were produced, while during growth on benzoate or 3-phenylpropionate crotonate served as a reducible co-substrate and was exclusively converted to butyrate. In the presence of crotonate interspecies hydrogen transfer was not necessary as a hydrogen sink. The benzoate degrader was isolated as a pure culture with crotonate as the only carbon source. The pure culture could also grow with benzoate/crotonate or 3-phenylpropionate/crotonate. The effect of high concentrations of crotonate and of acetate or butyrate on growth of the benzoate degrader was investigated. The benzoate degrader was compared with S. buswellii for its morphology, physiology and DNA base composition. Except for the fact that S. buswellii was also able to grow on cinnamate, no differences between the two organisms were detected. The isolate is named S. buswelli, strain GA.