Energetic aspects of CO oxidation in desulfovibrio desulfuricans

In cell suspension of Desulfovibrio desulfuricans B-1388, oxidation of CO as the only energy source is associated with reduction of SO42-. After a 2-h incubation of cells in 8% CO, 81% of the gas is converted. Oxidation of 1 mole CO results in formation of 0.23 mole H2S. Intracellular ATP content increases from 2.5 (control) to 8.3 nmoles/mg (during CO conversion). Dinitrophenol inhibits sulfate reduction and CO oxidation. CO dehydrogenase was detected in cytoplasmic and membrane cell fractions (59 and 34%, respectively).     

Periplasmic superoxide dismutase from Desulfovibrio desulfuricans 1388 is an iron protein

It is shown that the genome of the sulfate-reducing bacterium Desulfovibrio desulfuricans 1388 contains a superoxide dismutase (SOD) gene (sod). The gene encodes an export signal peptide characteristic for periplasmic redox proteins. The amino acid sequence showed high homology with iron-containing SODs from other bacteria. Electrophoretically pure SOD was isolated from the periplasmic fraction of bacterial cells by FPLC chromatography. Like other Fe-SODs, D. desulfuricans 1388 superoxide dismutase is inhibited by H2O2 and azide, but not by cyanide.     

Antioxidative enzymes of sulfate-reducing bacterium desulfovibrio desulfuricans: superoxide dismutase and peroxidases

Extracts of Desulfovibrio desulfuricans B-1388 cells grown under anaerobic conditions displayed superoxide dismutase activity. The maximal activity was found during the stationary growth phase. The enzyme was virtually completely located in the periplasm fraction. D. desulfuricans B-1388 lacked catalase activity but contained active NADH- and NADPH-peroxidases. The activity of NADH-peroxidase depended on the physiological state of the culture. On changing the growth conditions (the presence of 5% CO in the gaseous phase), the activity of superoxide dismutase decreased.     

Carbon monoxide and oxidative stress in Desulfovibrio desulfuricans B-1388

It has been shown that carbon monoxide (CO) in low concentration may be an active biochemical and physiological regulator of cell function. The bases of CO toxicity and cell protection are not clearly understood. To provide insights into these mechanisms, we measured superoxide production by D. desulfuricans B-1388 incubated anaerobically in Postgate medium with or without 5% CO. D. desulfuricans B-1388 growing with CO in the gas phase produced more superoxide radicals then control cells growing in Ar. When the cells were pregrown with CO, NADH oxidase and peroxidase activities were increased. The increase in peroxidase activities of cells growing under CO (particularly NADH peroxidase) suggested that H(2)O(2) was accumulated in cells. Superoxide dismutase (SOD) activity of cells decreased in exponential growth phase and increased in stationary phase. This may be due to CO concentration fall during CO oxidation by CO dehydrogenase. Altogether, our data suggest that superoxide production is a possible mechanism of CO toxicity.     

Biotechnological potential of sulfate-reducing bacteria for transformation of nitrocellulose

The biotransformation of NC by Desulfovibrio sp. was studied. The mass of NC was decreased by 4.9-9.3%. The rate of NC transformation was between 46 and 73 mg NC per mg of bacterial protein in 10 days. Moreover, N content (%N) in the remaining NC was reduced by 2-12%. The inhibitory effect of NC was clearly expressed when the growth of D. desulfuricans 1388 in lactate/sulfate medium was initiated. The growth rate of bacteria was 1.5-fold greater when NC was not added (0.074 and 0.05 h(-1) respectively). The transformation of NC by D. desulfuricans was accompanied by the appearance of nitrate in the culture liquid, the amount of which reached the peak by the 8th day.     

FTIR-spectroscopic studies of the fine structure of nitrocellulose treated by Desulfovibrio desulfuricans

Most studies have concluded that nitrocellulose (NC) with high degree of nitrogen content is resistant to biodegradation. Our results demonstrated that NC (>11%N) does undergo biotransformation in the presence of sulfate-reducing bacteria Desulfovibrio desulfuricans 1388. FTIR analyses indicated that the substitution of nitro groups for OH(-) groups took place. The spectrum of precipitate obtained after acetone extraction of NC resembled mainly the spectrum of native cellulose. Thus the synthetic unbiodegradable polymer was transformed to the natural compound accessible for microorganisms.     

Carbon monoxide inhibits superoxide dismutase and stimulates reactive oxygen species production by Desulfovibrio desulfuricans 1388

The hypothesis that oxidative stress characterized by enhanced superoxide generation underlies the toxicity of some factors to living organisms has been investigated. It is shown that CO (5-6% in gas phase) changed some growth parameters (mu, t(d)) of the sulfate-reducing bacterium Desulfovibrio desulfuricans 1388. Enhanced O(2)(-) generation registered by EPR spectroscopy and adrenochrome method was observed when cells were incubated under CO. The SOD activity in cells from the exponential growth phase growing under CO was decreased 1.5-fold compared with the control cells growing under Ar. SOD activities in cells from the stationary growth phase growing with or without CO were comparable. The results support the concept that CO toxicity for sulfate-reducing bacteria is an oxidative stress that arises in cells oxidizing CO to CO(2).     

Diversity and origin of Desulfovibrio species: phylogenetic definition of a family

The different nutritional properties of several Desulfovibrio desulfuricans strains suggest that either the strains are misclassified or there is a high degree of phenotypic diversity within the genus Desulfovibrio. The results of partial 16S rRNA and 23S rRNA sequence determinations demonstrated that Desulfovibrio desulfuricans ATCC 27774 and "Desulfovibrio multispirans" are closely related to the type strain (strain Essex 6) and that strains ATCC 7757, Norway 4, and El Agheila Z are not. Therefore, these latter three strains of Desulfovibrio desulfuricans are apparently misclassified. A comparative analysis of nearly complete 16S rRNA sequences in which we used a least-squares analysis method for evolutionary distances, an unweighted pair group method, a signature analysis method, and maximum parsimony was undertaken to further investigate the phylogeny of Desulfovibrio species. The species analyzed were resolved into two branches with origins deep within the delta subdivision of the purple photosynthetic bacteria. One branch contained five deep lineages, which were represented by (i) Desulfovibrio salexigens and Desulfovibrio desulfuricans El Agheila Z; (ii) Desulfovibrio africanus; (iii) Desulfovibrio desulfuricans ATCC 27774, Desulfomonas pigra, and Desulfovibrio vulgaris; (iv) Desulfovibrio gigas; and (v) Desulfomicrobium baculatus (Desulfovibrio baculatus) and Desulfovibrio desulfuricans Norway 4. A correlation between 16S rRNA sequence similarity and percentage of DNA relatedness showed that these five deep lineages are related at levels below the minimum genus level suggested by Johnson (in Bergey's Manual of Systematic Bacteriology, vol. 1, 1984). We propose that this branch should be grouped into a single family, the Desulfovibrionaceae. The other branch includes other genera of sulfate-reducing bacteria (e.g., Desulfobacter and Desulfococcus) and contains Desulfovibrio sapovorans and Desulfovibrio baarsii as separate, distantly related lineages.     

Hydrogen metabolism in Desulfovibrio desulfuricans strain New Jersey (NCIMB 8313)--comparative study with D. vulgaris and D. gigas species

This article aims to study hydrogen production/consumption in Desulfovibrio (D.) desulfuricans strain New Jersey, a sulfate reducer isolated from a medium undergoing active biocorrosion and to compare its hydrogen metabolism with two other Desulfovibrio species, D. gigas and D. vulgaris Hildenborough. Hydrogen production was followed during the growth of these three bacterial species under different growth conditions: no limitation of sulfate and lactate, sulfate limitation, lactate limitation, pyruvate/sulfate medium and in the presence of molybdate. Hydrogen production/consumption by D. desulfuricans shows a behavior similar to that of D. gigas but a different one from that of D. vulgaris, which produces higher quantities of hydrogen on lactate/sulfate medium. The three species are able to increase the hydrogen production when the sulfate became limiting. Moreover, in a pyruvate/sulfate medium hydrogen production was lower than on lactate/sulfate medium. Hydrogen production by D. desulfuricans in presence of molybdate is extremely high. Hydrogenases are key enzymes on production/consumption of hydrogen in sulfate reducing organisms. The specific activity, number and cellular localization of hydrogenases vary within the three Desulfovibrio species used in this work, which could explain the differences observed on hydrogen utilization.     

Mutual influences of Pseudomonas aeruginosa and Desulfovibrio desulfuricans on their adhesion to stainless steel

The mutual influences of Pseudomonas aeruginosa PAO1 and Desulfovibrio desulfuricans subsp. desulfuricans (ATCC 29577) on their adhesion to stainless steel were investigated in batch and column experiments. It was found that P. aeruginosa promoted the adhesion of D. desulfuricans under conditions of turbulence, but not under quiescent conditions. The enhancement involved the alignment of most D. desulfuricans along P. aeruginosa cells and was attributed to the additional interaction surface area provided by adhered P. aeruginosa to aligning D. desulfuricans cells. A slightly positive effect of preadhered D. desulfuricans on the adhesion of P. aeruginosa was found. Under condition of laminar flow, substantially better adhesion of D. desulfuricans to confluent P. aeruginosa biofilms than to steel was observed. The mutual influences are discussed in terms of more favorable adhesion energies and the influence of changed hydraulic conditions due to the roughness of P. aeruginosa biofilms.     

Characterization of periplasmic hydrogenase from Desulfovibrio vulgaris Miyazaki K

Periplasmic hydrogenase [hydrogen:ferricytochrome c3 oxidoreductase, EC 1.12.2.1] from Desulfovibrio vulgaris Miyazaki K (MK) was purified to homogeneity. Its chemical and immunological properties were examined and compared with those of other Desulfovibrio hydrogenases. The pure enzyme showed a specific activity of 1,000 mumol H2 evolution min-1 (mg protein)-1. The enzyme had a molecular weight of 50,000 as estimated by gel filtration and consisted of a single polypeptide chain. The absorption spectrum of the enzyme was characteristic of an iron-sulfur protein and the extinction coefficients at 400 and 280 nm were 34 and 104 mM-1. cm-1, respectively. It contained 9.4 mol iron and 6.9 mol of acid-labile sulfide per mol. The amino acid composition of the preparation was very similar to the value reported for D. desulfuricans NRC 49001 hydrogenase. Rabbit antisera were prepared against the enzyme of D. vulgaris MK. Ouchterlony double diffusion and immunotitration tests of crude extracts from several strains of Desulfovibrio revealed that the enzyme from MK cells was immunologically identical with those from D. vulgaris Hildenborough and D. desulfuricans NRC 49001, but different from those from D. vulgaris Miyazaki F (MF) and Miyazaki Y, and D. desulfuricans Essex 6 strains. It is concluded that among Desulfovibrio hydrogenases, those from D. vulgaris MK, D. vulgaris Hildenborough and D. desulfuricans NRC 49001 form one group in terms of both subunit structure and antigenicity.     

In vivo 31P-NMR studies of Desulfovibrio species. Detection of a novel phosphorus-containing compound

The phosphorus metabolism of sulfate-reducing bacteria was, for the first time, probed by in vivo 31P NMR. A novel phosphoric anhydride diester compound was detected in Desulfovibrio desulfuricans ATCC 27774 at intracellular concentrations up to 5 mM. The compound has been extracted and partially purified by anion-exchange chromatography and analysed by 31P, 13C and 1H NMR. These studies show that the novel phosphorus-containing compound is formed by five carbon atoms and is probably cyclic, with a Mr of approximately 300. Various Desulfovibrio strains were examined in vivo for the presence of this phosphorus-containing compound. Detectable amounts of the novel metabolite were found in D. desulfuricans ATCC 27774 when grown on lactate/sulfate, lactate/thiosulfate or pyruvate/sulfate. The phosphorus-containing compound was not detected when this strain of D. desulfuricans was grown on lactate/nitrate or pyruvate; neither was it detected in two other strains which, like D. desulfuricans ATCC 27774, have the capability of utilizing nitrate as a terminal electron acceptor.     

Purification and characterization of cytochrome c3, ferredoxin, and rubredoxin isolated from Desulfovibrio desulfuricans Norway.

Different electron carriers of the non-desulfoviridin-containing, sulfate-reducing bacterium Desulfovibrio desulfuricans (Norway strain) have been studied. Two nonheme iron proteins, ferredoxin and rubredoxin, have been purified. This ferredoxin contains four atoms of non-heme iron and acid-labile sulfur and six residues of cysteine per molecule. Its amino acid composition suggests that it is homologous with the other Desulfovibrio ferredoxins. The rubredoxin is also an acidic protein of 6,000 molecular weight and contains one atom of iron and four cysteine residues per molecule. The amino acid composition and molecular weight of the cytochrome c3 from D. desulfuricans (strain Norway 4) are reported. Its spectral properties are very similar to those of the other cytochromes c3 (molecular weight, 13,000) of Desulfovibrio and show that it contains four hemes per molecule. This cytochrome has a very low redox potential and acts as a carrier in the coupling of hydrogenase and thiosulfate reductase in extracts of Desulfovibrio gigas and Desulfovibrio desulfuricans (Norway strain) in contrast to D. gigas cytochrome c3 (molecular weight, 13,000). A comparison of the activities of the cytochrome c3 (molecular weight, 13,000) of D. gigas and that of D. desulfuricans in this reaction suggests that these homologous proteins can have different specificity in the electron transfer chain of these bacteria.     

Biochemical and spectroscopic characterization of an aldehyde oxidoreductase isolated from Desulfovibrio aminophilus

Aldehyde oxidoreductase (AOR) activity has been found in a number of sulfate-reducing bacteria. The enzyme that is responsible for the conversion of aldehydes to carboxylic acids is a mononuclear molybdenum enzyme belonging to the xanthine oxidase family. We report here the purification and characterization of AOR isolated from the sulfate-reducing bacterium Desulfovibrio (D.) aminophilus DSM 12254, an aminolytic strain performing thiosulfate dismutation. The enzyme is a homodimer (ca. 200 kDa), containing a molybdenum centre and two [2Fe-2S] clusters per monomer. UV/Visible and electron paramagnetic resonance (EPR) spectra of D. aminophilus AOR recorded in as-prepared and reduced states are similar to those obtained in AORs from Desulfovibrio gigas, Desulfovibrio desulfuricans and Desulfovibrio alaskensis. Despite AOR from D. aminophilus is closely related to other AORs, it presents lower activity towards aldehydes and no activity towards N-heterocyclic compounds, which suggests another possible role for this enzyme in vivo. A comparison of the molecular and EPR properties of AORs from different Desulfovibrio species is also included.     

Isolation of a New Pigment, Desulforubidin, from Desulfovibrio desulfuricans (Norway Strain) and Its Role in Sulfite Reduction

A new pigment, desulforubidin, that has sulfite reducing activity, has been purified from extracts of the Norway strain of Desulfovibrio desulfuricans, which lacks desulfoviridin.     

Possible involvement of a L-delta 1-pyrroline-5-carboxylate (P5C) reductase in the synthesis of proline in Desulfovibrio desulfuricans Norway

A L-delta 1-pyrroline-5-carboxylate reductase activity has been detected in crude extracts of Desulfovibrio desulfuricans Norway. This P5C reductase activity is also found when a 2.5 kb D. desulfuricans DNA fragment is introduced into an Escherichia coli proC mutant. Although it restores growth of the proC mutant, the ProDd enzyme might be detrimental to the E. coli host since the plasmid carrying the cognate proDd gene is segregated at high rate by the cells but is stabilized by small deletions which lead to a loss of the P5C reductase activity.     

Pyruvate-carbon dioxide exchange reaction of Desulfovibrio desulfuricans

Suh, Byungse (University of Kansas, Lawrence), and J. M. Akagi. Pyruvate-carbon dioxide exchange reaction of Desulfovibrio desulfuricans. J. Bacteriol. 91:2281-2285. 1966.-The pyruvate-CO(2) exchange reaction, catalyzed by Desulfovibrio desulfuricans, required the presence of phosphate and coenzyme A. However, the requirement for phosphate disappeared when the concentration of coenzyme A was increased to a level of 3.8 x 10(-3)m. Passing crude extracts through a diethylaminoethyl-cellulose column and an Amberlite CG-50 ion-exchange column, to remove ferredoxin and cytochrome c(3), resulted in a marked decrease in exchange activity; full activity was restored by the addition of ferredoxin or cytochrome c(3). Fe(++) or Co(++) stimulated the exchange of CO(2) into pyruvate.     

Induction and partial purification of bacteriophages from Desulfovibrio vulgaris (Hildenborough) and Desulfovibrio desulfuricans ATCC 13541.

Bacteriophages were induced from cultures of Desulfovibrio vulgaris NCIMB 8303 and Desulfovibrio desulfuricans ATCC 13541 by UV light. The optimum time of UV exposure was 1 min and the maximum yield of phage was obtained 9-10 h after UV treatment. The two phage preparations were compared by restriction enzyme analysis and Southern blot hybridization. The nucleic acid from both phages was cut by restriction endonucleases specific for double-stranded DNA. The phage DNAs from D. vulgaris and D. desulfuricans showed different restriction enzyme cleavage patterns. No homology was observed between a 25 kb probe from the D. vulgaris phage DNA and the phage DNA from D. desulfuricans. Protein profiles of the phages from both sources were also studied; the D. vulgaris phage contained two major bands corresponding to Mr values of 37 000 and 56 000 while the D. desulfuricans phage contained only one major band, of Mr 38 000.