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I N Gogotov

Publications and source records attributed to I N Gogotov.

At least 19 recordsLinked to original sources
PubMed2004

Properties of stable hydrogenase from the purple sulfur bacterium Lamprobacter modestohalophilus.

Some properties of a hydrogenase from the recently isolated phototrophic sulfur bacterium Lamprobacter modestohalophilus strain Syvash and its resistance to a number of inactivating factors have been investigated. The enzyme consists of two subunits, 64 and 30 kD; pI = 4.5. The optimal pH was 8.5-9.5 for hydrogen uptake and 4.0 for H2 evolution. Hydrogenase preparations were resistant to the effects of O2, CO, and temperature, revealing high stability under storage. A considerable inactivation of the enzyme was observed at temperatures above 80 degrees C; the temperature optimum of methyl viologen reduction by H2 was 85 degrees C. Inhibitory effects of Ni2+, Cd2+, and Mg2+ on the hydrogenase activity were shown to be reversible and competitive with respect to methyl viologen in the hydrogen oxidation reaction.

Bacterial Proteins↗
PubMed2000

Influence of metal ions on hydrogenase from the purple sulfur bacterium Thiocapsa roseopersicina.

The effects of some metal ions on the activity and activation of Thiocapsa roseopersicina hydrogenase have been studied. Inhibitory effects of Ni2+ and Cd2+ on the catalytic activity of the enzyme were reversible and competitive with respect to methyl viologen (MV) in the reaction of hydrogen oxidation. The affinity of these metal ions to the enzyme increased significantly with increasing pH, suggesting that their interactions are determined by electrostatic forces. Cu2+ and Hg2+ irreversibly inhibited the hydrogenase activity. A decrease in absorption of hydrogenase at 400 nm in the presence of these metal ions is indicative of the destruction of the FeS cluster in the enzyme.

Catalysis↗
PubMed1996

Reversible hydrogenase of Anabaena variabilis ATCC 29413: catalytic properties and characterization of redox centres.

The catalytic and spectroscopic properties of the reversible hydrogenase from the cyanobacterium Anabaena variabilis have been examined. The hydrogenase required reductive activation in order to elicit hydrogen-oxidation activity. Carbon monoxide was a weak (Ki=35 microM), reversible and competitive inhibitor. A flavin with the chromatographic properties of FMN, and nickel were detected in the purified enzyme. A. variabilis hydrogenase exhibited electron paramagnetic resonance (EPR) spectra in its hydrogen-reduced state, indicative of [2Fe-2S] and [4Fe-4S] clusters. Although no EPR signals due to nickel were detected, the results are consistent with the enzyme being a flavin-containing hydrogenase of the nickel-iron type.

Anabaena↗
PubMed1993

Purification and properties of a flavodoxin from the heterocystous cyanobacterium Anabaena sphaerica.

A flavodoxin was purified to homogeneity from the nitrogen-fixing heterocystous cyanobacterium Anabaena sphaerica grown under iron-limited conditions. The protein has a molecular mass of 21 kDa, and its spectral properties and amino-acid composition are very close to that of flavodoxins from other cyanobacteria. A. sphaerica flavodoxin supported the activities of A. sphaerica NADP reductase and Clostridium butyricum hydrogenase in reconstituted systems with illuminated plant chloroplasts as reductant. With the use of polyclonal anti-flavodoxin antiserum it was found that nitrogen-fixing cultures of A. sphaerica grown under iron-sufficient conditions contain low but significant amounts of flavodoxin (0.2-0.6 micrograms/mg crude extract protein) which increased dramatically (to 8-15 micrograms/mg crude extract protein) after the iron concentration in the medium was decreased to below 1 microM Fe. The flavodoxin content of both iron-limited and iron-sufficient. A. sphaerica was also shown to depend upon the growth phase of the (batch) cultures with a maximum at early exponential phase, coinciding with maximal in-vivo nitrogenase activity. These results suggest that A. sphaerica flavodoxin not only substitutes for ferredoxin under iron-limiting conditions, but also fulfills some specific role under iron-sufficient conditions.

Amino Acids↗
PubMed1993

Purification and properties of a bacterial-type ferredoxin from the nitrogen-fixing cyanobacterium Anabaena variabilis ATCC29413.

Three soluble ferredoxins were purified to homogeneity from nitrogen-fixing cultures of Anabaena variabilis (ATCC 29413) and characterized. The purified proteins have different absorption spectra, molecular mass, iron content, amino-acid composition and resistance to O2 inactivation. Two were plant-type ferredoxins FdI and FdxH, corresponding to the previously reported ferredoxins II and I (Böhme, H. and Schrautemeier, B. (1987) Biochim. Biophys. Acta 891, 1-7). The third ferredoxin (ferredoxin III) (previously not described in cyanobacteria) was a bacterial-type ferredoxin. Ferredoxin III has a molecular mass of about 6 kDa and contains 3-4 atoms Fe/mol. Native (oxidized) ferredoxin III shows an EPR-signal at g = 2.014 that disappears after reduction by dithionite, characteristic of ferredoxins containing three-iron clusters. Ferredoxin III, like ferredoxin FdxH, is inactivated by oxygen. Ferredoxin III supports higher rates of C2H2 reduction by Rhodobacter capsulatus nitrogenase than FdI and higher rates of H2 evolution by clostridial hydrogenase than FdI and FdxH. Combined nitrogen suppresses the synthesis of both nitrogenase and ferredoxin III. These data suggest a possible role of ferredoxin III (bacterial-type) in nitrogen fixation by A. variabilis.

Amino Acids↗
PubMed1986

Hydrogenases of phototrophic microorganisms.

This review surveys recent work done in the laboratory of the author and related laboratories on the properties and possible practical applications of hydrogenases of phototrophic microorganisms. Homogeneous hydrogenase preparations were obtained from purple non-sulfur (Rhodospirillum rubrum S1, Rhodobacter capsulatus B10) and purple sulfur (Chromatium vinosum D, Thiocapsa roseopersicina BBS) bacteria, and from the green sulfur bacterium Chlorobium limicola forma thiosulfatophilum L; highly purified hydrogenase samples were prepared from the cyanobacterium Anabaena cylindrica and from the green alga Chlamydomonas reinhardii. It was shown that hydrogenases of R. capsulatus and T. roseopersicina contain Ni and Fe-S cluster. The cytochromes of the c or b type serve as native electron acceptors for the hydrogenases of the purple bacteria and cyanobacteria; rubredoxin or cytochrome c for the hydrogenase of the green sulfur bacterium; and ferredoxin for Ch. reinhardii hydrogenase. The hydrogenase of T. roseopersicina BBS reversibly activates H2 at Eh less than -290 mV (pH 7), whereas those from R. capsulatus and from C. limicola f. thiosulfatophilum exhibit their maximum activity at Eh greater than -300 mV and are thus favourable only for the H2 uptake. Hydrogenase synthesis in different phototrophs depends on pO2, H2 concentrations and organic substrates. Organic compounds, which serve as electron donors and carbon sources, repress hydrogenase synthesis in R. rubrum, R. capsulatus and in Ectothiorhodospira shaposhnikovii when present at high concentrations. The synthesis of T. roseopersicina hydrogenase is constitutive. H2 notably stimulates hydrogenase activity in R. capsulatus. The synthesis of hydrogenase in R. sphaeroides 2R occurs only in the presence of H2 and does not depend on the presence of organic compounds in the medium.

Bacteria↗
PubMed1983

[Properties of two forms of ferredoxin from Rhodopseudomonas capsulata].

Electrophoretically homogenous preparations of two forms of ferredoxin were isolated from the nitrogen-fixing cells of the purple non-sulphur bacterium Rhodopseudomonas capsulata B10. The values of Mr for ferredoxins I and II are 12000 and 18000, respectively. Ferredoxin I contains 8 atoms of Fe+2 and 8 atoms of S2-; ferredoxin II--4 atoms of Fe2+ and 4 atoms of S2- per molecule. The ferredoxins differ also in their absorption spectra, stability and catalytic activity during electron transfer to nitrogenase of Rh. capsulata. The reduction of C2H2 and evolution of H2 in the presence of ferredoxin I occurs twice as fast as compared to that in the presence of ferredoxin II. Ferredoxin I synthesis takes place in the nitrogen-fixing cells of Rh. capsulata alone, whereas ferredoxin II formation does not depend on the growth conditions.

Electron Transport↗
PubMed1983

[High potential iron-sulfur protein from Thiocapsa roseopersicina].

A high potential protein (HiPIP) containing a [4Fe-4S] cluster was obtained in a homogeneous state from the purple sulfur bacterium T. roseopersicina strain BBS. The EPR and absorption spectra are specific for this type of proteins. In the absorption spectrum the A283/A390 ratio equals to 2.25; Mr = 10 000, pI is 4.08, E0' = +328 mV. The reduced form of HiPIP was found to be more stable upon storage; T 1/2 is 40 min at 80 degrees C. HiPIP can be reduced by ascorbate, cysteine, 2-mercaptoethanol and sulfide as well as by formate, H2 and NADPH in the presence of corresponding enzymes. Chromatophores oxidize HiPIP in the light. This suggests that the latter takes part in the photosynthetic electron transfer chain of T. roseopersicina.

Chromatium↗
PubMed1982

[Low potential c-type cytochrome of Thiocapsa roseopersicina].

The low potential c-type cytochrome from the phototrophic purple sulphur bacterium Thiocapsa roseopersicina, strain BBS was isolated in electrophoretically homogeneous state. The bulk of the cytochrome (approximately 90%) after disruption of the cells remained in the membrane fraction. The absorption spectrum of the cytochrome was characterized by the maxima at 420, 523 and 552 nm in the reduced state and at 408 nm in the oxidized one. The cytochrome interacted with CO in the reduced state. The molecular weight of the cytochrome is 50 000. The cytochrome contains great amounts of phenylalanine, leucine, valine, aspartic and glutamic acids and can be reduced by dithionite but not by cysteine, sulfide or ascorbate. Besides, the cytochrome can also be reduced by NAD(P)H in the presence of NAD(P)-reductases of T. roseopersicina, when ferredoxin of Spirulina platensis or benzyl viologen are added to the reaction mixture. The cytochrome can act as an electron donor (acceptor) for T. roseopersicina hydrogenase.

Carbon Monoxide↗
PubMed1982

[Stability of hydrogenase from the purple sulfur bacteria Thiocapsa roseopersicina].

The hydrogenase from T. roseopersicina is highly resistant to the effects of urea (8 M), Me2SO (20%) and DS-Na (1%), while inactivation of the hydrogenase from Rhodopseudomonas capsulata occurs in the presence of 0.1% DS-Na. The higher the purification level of T. roseopersicina hydrogenase preparation, the higher stability it possesses (T 1/2 = 60 days, 24 degrees). The hydrogenase inactivation at 80 degrees under anaerobic conditions occurs in one stage according to the equation of first-order-reaction (k1i = 7.10(-5) sec-1), while under aerobic conditions it has two stages with a decrease in the rate of this process in the second stage (k2i = 1.8 . 10(-6) sec-1). Glycerol and NaCl do not stabilize the T. roseopersicina hydrogenase. The rate of thermal inactivation of the hydrogenase bound to the membranes, DEAE-cellulose or phenylsepharose is higher than that of the soluble enzyme. The considerable decrease of the thermal stability of the enzyme is caused by the thiol reagents: they cause irreversible denaturation of the enzyme. The hydrogenase partly inactivated under aerobic conditions is reactivated in the presence of Na2S2O4. The data obtained indicate the important role of disulphide bonds in stabilization of T. roseopersicina hydrogenase.

Aerobiosis↗
PubMed1980

[Purification and properties of cytochrome c552 from purple sulfur bacterium Thiocapsa roseopersicina].

The method of purification up to electrophoretical homogeneity of cytochrome c552 from the phototrophic bacterium Thiocapsa roseopersicina, strain BBS is described. For the cytochrome absorption spectrum the maxima at 417, 523 and 552 nm are characteristic for the reduced state and at 409 nm--for the oxidized state. The molecular weight is equal to 62000. The cytochrome contains two hemes per molecule and consists of two subunits. pI is 4.1; E0' is about 10 mV. Cytochrome c552 is a flavoprotein according to its fluorescence spectrum and subunit structure. T. roseopersicina cytochrome c552 is able to be reduced with sulphide, cysteine and ascorbate as well as with H2 in the presence of hydrogenase from the same bacterium. These data suggest that cytochrome c552 from T. roseopersicina functions in vivo at the initial stage of electron transport from hydrogen and sulphide.

Chromatiaceae↗
PubMed1980

[Purification and properties of NAD-reductase from phototrophic bacterium Thiocapsa roseopersicina].

The purification by affinity chromatography up to homogeneity and the properties of NAD-reductase from purple sulfur bacterium Thiocapsa roseopersicina, strain BBS, are described. The molecular weight of NAD-reductase is about 80000; pI is 3.9. The enzyme consists of two subunits. According to the stabilizing effect of FAD at preparative electrophoresis and the inhibitory effect of atebrine NAD-reductase is a flavoprotein. The bulk of the enzyme (about 75%) is localized in the cell periplasmic space. NAD-reductase is less thermostable and has a lower O2 stability as compared to the NADP-reductase from the same organism. The enzyme is specific to NADH ane catalyzes the menadione-reductase reaction, diaphorase reaction of benzyl viologen and methyl viologen reductions. In the presence of NADH NAD-reductase reduces cytochromes c552 and "c3" from T. roseopersicina and forms a complex with spinach ferredoxin.

Chromatiaceae↗
PubMed1978

Superoxide dismutase and catalase in the protection of the proton-donating systems of nitrogen fixation in the blue-green alga Anabaena cylindrica.

1. Superoxide dismutase activity was present in the heterocysts and vegetative cells of Anabaena cylindrica, but was always lower in the heterocysts. 2. No qualitative differences were found in the superoxide dismutase from the two cellular types. 3. Catalase activity was also present in both cellular types. 4. Most of the NADP reductase activity, as assayed with menadione or ferredoxin as electron acceptor, was localized within the heterocysts. 5. Studies on H2 consumption showed that most of the hydrogenase activity was associated with the heterocysts. 6. The results are discussed in terms of the postulate that superoxide dismutase and catalase are involved in the protection of the proton-donating systems participating in N2 fixation and H2 metabolism of heterocysts.

Catalase↗
PubMed1978

The properties of hydrogenase from Thiocapsa roseopersicina.

The soluble and chromatophore-bound hydrogenases from the purple sulphur bacterium Thiocapsa roseopersicina strain BBS were purified up to homogeneity and the properties studied. The amino acid composition of hydrogenase preparations from different fractions of T. roseopersicina is identical, glycine and alanine as N-terminal amino acid residues. In comparison with other hydrogenases, especially in the immobilized state, the preparations obtained are shown to be more stable to O2 during storage and they are characterized by high thermal stability. Inactivation is observed above 78--80 degrees C and the optimal temperature for enzyme action is 70 degrees C. The homogeneous enzyme preparations catalyse the exchange reaction between 2H2 and H2O and reversible redox reactions of methyl viologen and benzyl viologen as well as H2 formation from reduced ferredoxin. According to our data, the hydrogenase of T. roseopersicina bound with chromatophores is identical to the soluble one.

Amino Acids↗
PubMed1978

Relationships in hydrogen metabolism between hydrogenase and nitrogenase in phototrophic bacteria.

Purple bacteria Rhodospirillum rubrum and Thiocapsa roseopersicina form two enzymes, hydrogenase and nitrogenase, which participate in hydrogen metabolism. H2 photoproduction in these bacteria is associated mainly or completely with the action of nitrogenase. The soluble and membrane-bound hydrogenases of T. roseopersicina have similar physicochemical properties (mol. weight, subunit composition, N-terminal amino acids, Fe2+ and S2- content, pl. Eo'). In comparison with other hydrogenases the enzyme from R. rubrum and T. roseopersicina evolve H2 with high rate from reduced cytochrome c3, but not from ferredoxins. H2 production and N2 fixation take place in the presence of NAD(P)H. NADP-reductase, ferredoxin and cytochrome c3 participate in this reaction. Possible relationships between hydrogenase-nitrogenase in the metabolism of molecular hydrogen are discussed.

Bacteria↗
PubMed1978

Hydrogen evolution by chloroplast-hydrogenase systems: improvements and additional observations.

An in vitro system containing isolated chloroplasts, ferredoxin and bacterial hydrogenase on illumination evolves H2 and O2 from water. Maximum rate of hydrogen production so far achieved is two litres H2 per g. chlorophyll per h. The rate of H2 evolution per mg chlorophyll is dependent on concentrations of chlorophyll and ferredoxin in the reaction mixture. The rates as well as duration of H2 production are enhanced by the presence of oxygen scavengers and bovine serum albumin in the system. Hydrogenases and ferredoxins vary in their degree of cross reactivity in the chloroplast system; with some hydrogenases the H2 evolution rates were increased by the presence of additional biological electron carriers. Attempts to couple algal hydrogenases to the chloroplasts system have not succeeded so far.

Bacteria↗
PubMed1977

[Purification and properties of phototrophic bacteria Thiocapsa roseopersicina hydrogenase bound with chromatophores].

The method of solution and puridication of hydrogenase from chromatophores of purpur sulphur bacteria Thiocapsa roseopersicina strain BBS are described. Hydrogenase molecular weight is 73000. It contains 4,4 mole S2- and 3.1 mole Fe2+ per mole of protein; pI 4.15. The enzyme absorption spectrum has the maximun et 400-410 nm, which is characteristic of proteins containing non-haem iron. Membrane--linked enzyme as well as soluble hydrogenase of that microorganism is characterized by high thermal stability: inactivation occurs at the temperature above 78 degrees C when the optimal temperature for that enzyme is 70 degrees C. Homogenous enzyme catalyses D2--H2O exchange reaction, reversible redox reaction of methyl viologene and benzyl viologene.

Bacterial Chromatophores↗