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.     

Elemental sulfur as an intermediate of sulfide oxidation with oxygen byDesulfobulbus propionicus

The sulfate-reducing bacteriumDesulfobulbus propionicus oxidized sulfide, elemental sulfur, and sulfite to sulfate with oxygen as electron acceptor. Thiosulfate was reduced and disproportionated exclusively under anoxic conditions. When small pulses of oxygen were added to washed cells in sulfide-containing assays, up to 3 sulfide molecules per O₂ disappeared transiently. After complete oxygen consumption, part of the sulfide reappeared. The intermediate formed was identified as elemental sulfur by chemical analysis and turbidity measurements. When excess sulfide was present, sulfur dissolved as polysulfide. This process was faster in the presence of cells than in their absence. The formation of sulfide after complete oxygen consumption was due to a disproportionation of elemental sulfur (or polysulfide) to sulfide and sulfate. The uncoupler tetrachlorosalicylanilide (TCS) and the electron transport inhibitor myxothiazol inhibited sulfide oxidation to sulfate and caused accumulation of sulfur. In the presence of the electron transport inhibitor 2-n-heptyl-4-hydroxyquinoline-N-oxide (HQNO), sulfite and thiosulfate were formed. During sulfur oxidation at low oxygen concentrations, intermediary formation of sulfide was observed, indicating disproportionation of sulfur also under these conditions. It is concluded that sulfide oxidation inD. propionicus proceeds via oxidation to elemental sulfur, followed by sulfur disproportionation to sulfide and sulfate. Dedicated to Prof. Dr. Dr. h.c. Norbert Pfennig on the occasion of his 70th birthday     

Microbial acceleration of aerobic pyrite oxidation at circumneutral pH

Pyrite (FeS₂) is the most abundant sulfide mineral on Earth and represents a significant reservoir of reduced iron and sulfur both today and in the geologic past. In modern environments, oxidative transformations of pyrite and other metal sulfides play a key role in terrestrial element partitioning with broad impacts to contaminant mobility and the formation of acid mine drainage systems. Although the role of aerobic micro‐organisms in pyrite oxidation under acidic‐pH conditions is well known, to date there is very little known about the capacity for aerobic micro‐organisms to oxidize pyrite at circumneutral pH. Here, we describe two enrichment cultures, obtained from pyrite‐bearing subsurface sediments, that were capable of sustained cell growth linked to pyrite oxidation and sulfate generation at neutral pH. The cultures were dominated by two Rhizobiales species (Bradyrhizobium sp. and Mesorhizobium sp.) and a Ralstonia species. Shotgun metagenomic sequencing and genome reconstruction indicated the presence of Fe and S oxidation pathways in these organisms, and the presence of a complete Calvin–Benson–Bassham CO₂ fixation system in the Bradyrhizobium sp. Oxidation of pyrite resulted in thin (30–50 nm) coatings of amorphous Fe(III) oxide on the pyrite surface, with no other secondary Fe or S phases detected by electron microscopy or X‐ray absorption spectroscopy. Rates of microbial pyrite oxidation were approximately one order of magnitude higher than abiotic rates. These results demonstrate the ability of aerobic microbial activity to accelerate pyrite oxidation and expand the potential contribution of micro‐organisms to continental sulfide mineral weathering around the time of the Great Oxidation Event to include neutral‐pH environments. In addition, our findings have direct implications for the geochemistry of modern sedimentary environments, including stimulation of the early stages of acid mine drainage formation and mobilization of pyrite‐associated metals.     

Siroheme sulfite reductase isolated from Chromatium vinosum. Purification and investigation of some of its molecular and catalytic properties

Cells of the phototrophic bacterium Chromatium vinosum strain D were shown to contain a siroheme sulfite reductase after autotrophic growth in a sulfide/bicarbonate medium. The enzyme could not be detected in cells grown heterotrophically in a malate/sulfate medium. Siroheme sulfite reductase was isolated from autotrophic cells and obtained in an about 80% pure preparation which was used to investigate some molecular and catalytic properties of the enzyme. It was shown to consist of two different types of subunits with molecular weights of 37,000 and 42,000, most probably arranged in an ₄₄-structure. The molecular weight of the native enzyme was determined to 280,000, 51 atoms of iron and 47 atoms of acid-labile sulfur were found per enzyme molecule. The absorption spectrum indicated siroheme as prosthetic group; it had maxima at 280 nm, 392 nm, 595 nm, and 724 nm. The molar extinction coefficients were determined as 302×10³ cm²xmmol^-1 at 392 nm, 98×10³ cm² xmmol^-1 at 595 nm and 22×10³ cm²x-mmol^-1 at 724 nm. With reduced viologen dyes as electron donor the enzyme reduced sulfite to sulfide, thiosulfate, and trithionate. The turnover number with 59 (2 e^-/enzyme moleculexmin) was low. The pH-optimum was at 6.0. C. vinosum sulfite reductase closely resembled the corresponding enzyme from Thiobacillus denitrificans and also desulfoviridin, the dismilatory sulfite reductase from Desulfovibrio species. It is proposed that C. vinosum catalyses anaerobic oxidation of sulfide and/or elemental sulfur to sulfite in the course of dissimilatory oxidation of reduced sulfur compounds to sulfate.Non-common abbreviations APS adenylyl sulfate - SDS sodium dodecyl sulfate     

Utilization of sulfide and elemental sulfur by Ectothiorhodospira halochloris

Ectothiorhodospira halochloris grows photoheterotrophically with a variety of sulfur sources. During sulfide oxidation to elemental sulfur considerable amounts of polysulfides may be accumulated transiently. When grown on elemental sulfur no sulfate was produced by oxidation, but sulfide and polysulfide were formed by reduction. Only one soluble cytochrome c-551 was isolated and purified. It was a small acidic hemeprotein with a molecular weight of 6,300, an isoelectric point of 3.1 and a redox potential of-11 mV at pH 7.0. It showed three absorption maxima in the reduced state (=551 nm; =523 nm; =417 nm). The addition of various c-type cytochromes to a suspension of spheroplasts stimulated the velocity of sulfide oxidation. This stimulation was best with the small acidic cytochromes from E. halochloris or Ectothiorhodospira abdelmalekii. Sulfide oxidation was stopped by several uncoupling agents, ionophores and electron transport inhibitors. Antimycin A, rotenone and cyanide had no effect on sulfide oxidation.Dedicated to Prof. Dr. H. G. Schlegel on the occasion of his 60th birthday     

Oxidation of H2, organic compounds and inorganic sulfur compounds coupled to reduction of O2 or nitrate by sulfate-reducing bacteria

All of fourteen sulfate-reducing bacteria tested were able to carry out aerobic respiration with at least one of the following electron donors: H₂, lactate, pyruvate, formate, acetate, butyrate, ethanol, sulfide, thiosulfate, sulfite. Generally, we did not obtain growth with O₂ as electron acceptor. The bacteria were microaerophilic, since the respiration rates increased with decreasing O₂ concentrations or ceased after repeated O₂ additions. The amounts of O₂ consumed indicated that the organic substrates were oxidized incompletely to acetate; only Desulfobacter postgatei oxidized acetate with O₂ completely to CO₂. Many of the strains oxidized sulfite (completely to sulfate) or sulfide (incompletely, except Desulfobulbus propionicus); thiosulfate was oxidized only by strains of Desulfovibrio desulfuricans; trithionate and tetrathionate were not oxidized by any of the strains. With Desulfovibrio desulfuricans CSN and Desulfobulbus propionicus the oxidation of inorganic sulfur compounds was characterized in detail. D. desulfuricans formed sulfate during oxidation of sulfite, thiosulfate or elemental sulfur prepared from polysulfide. D. propionicus oxidized sulfite and sulfide to sulfate, and elemental sulfur mainly to thiosulfate. A novel pathway that couples the sulfur and nitrogen cycles was detected: D. desulfuricans and (only with nitrite) D. propionicus were able to completely oxidize sulfide coupled to the reduction of nitrate or nitrite to ammonia. Cell-free extracts of both strains did not oxidize sulfide or thiosulfate, but formed ATP during oxidation of sulfite (37 nmol per 100 nmol sulfite). This, and the effects of AMP, pyrophosphate and molybdate on sulfite oxidation, suggested that sulfate is formed via the (reversed) sulfate activation pathway (involving APS reductase and ATP sulfurylase). Thiosulfate oxidation with O₂ probably required a reductive first step, since it was obtained only with energized intact cells.Abbreviations CCCP carbonyl cyanide m-chlorophenylhydrazone - APS adenosine phosphosulfate or adenylyl sulfate     

Coexistence of Chlorobium and Chromatium in a sulfide-limited continuous culture

Competition experiments between Chromatium vinosum and Chlorobium limicola in sulfide-limited continuous culture under photolithoautotrophic conditions resulted in the coexistence of both organisms. The ratio between the two bacteria was dilution-rate as well as pH dependent. The observed coexistence can be explained as a hitherto not reported form of dual substrate limitation. The two substrates involved are the electron donors sulfide (growth-limiting substrate in the reservoir vessel) and extracellular elemental sulfur (formed by Chlorobium as a result of sulfide oxidation). It is argued that, although Chlorobium may have the better affinity for both substrates involved, Chromatium can compete successfully on the basis of its intracellular storage of sulfur. Ecological implication of the observed coexistence with respect to natural blooms are discussed.     

Sulfur Transformation in Microbially Mediated Pyrite Oxidation by Acidithiobacillus ferrooxidans: Insights From X-ray Photoelectron Spectroscopy-Based Quantitative Depth Profiling

The oxidation of pyrite and other sulfides is responsible for the generation of acid mine drainage and acid rock drainage, which leads to further contamination of soil and water. In these processes, microbial oxidation usually prevails over chemical oxidation. To determine the mechanism of microbial oxidation of pyrite, the interaction of Acidithiobacillus ferrooxidans with pyrite was comprehensively studied, and the sulfur transformation in the interaction was disclosed using X-ray photoelectron spectroscopy (XPS) depth profiling. Abundant bacterial cells attach to pyrite surface and form biofilms, which greatly enhances surface corrosion and results in two types of etching pits: bacteria-driven rod-shaped and chemically driven hexagonal etching pits. The details of XPS depth profiles on a reacted pyrite surface reveal that the surface sulfur was first oxidized into elemental sulfur. Thereafter, elemental sulfur was further oxidized to intermediate species S₂O₃^2?, SO₃^2?, and ultimately to SO₄^2?. The oxidation sequence of sulfur is S₂^2?/S^2?→S_n^2?, S⁰→SO₃^2?, and S₂O₃^2?→SO₄^2?. Meanwhile, the remnant ferrous iron in the surface layer was released into solution and subsequently oxidized into Fe³⁺ by A. ferrooxidans and dissolved oxygen, which in turn enhanced the oxidation of sulfur. Fe³⁺, sulfate, and other ions (e.g., K⁺, Na⁺, NH₄⁺) in the solution precipitated as jarosite, hydroniumjarosite, and ammoniojarosite. On the basis of results, a three-staged model is proposed to interpret the kinetics of microbial oxidation of pyrite.     

Nitrogen fixation activity of a filamentous,nonheterocystous cyanobacterium in the presence and absence of exogenous,organic substrates

Gallionella ferruginea is able to utilize Fe(II) and the reduced sulfur compounds sulfide and thiosulfate as electron donor and energy source. Tetrathionate and elemental sulfur, on the other hand, are not metabolized. In sulfide-O₂ microgradient cultures G. ferruginea grows at the interface between the oxidizing and the reducing zones. Optimal growth depends on low oxygen and sulfide concentrations. Establishing within the gradient protects the bacterium from too high sulfide concentrations. G. ferruginea excretes extracellular polymeric substances (EPS). While in FeS-gradient cultures 2×10⁶ cells/ml were obtained the bacterial mass could be increased to 1–3×10⁸ cells/ml in shaken batch cultures using thiosulfate as substrate. A further increase of bacterial mass by adding an organic carbon source was not possible confirming that G. ferruginea is an obligate autotrophic organism. When growing on sulfide or thiosulfate the otherwise characteristic twisted stalk consisting of ferric hydroxide is lacking. It is thus shown to be a metabolic end product of Fe(II) oxidation rather than metabolically active cellular material.     

Protein synthesis by Beggiatoa alba B18LD in the presence and absence of sulfide

The interaction of sulfide oxidation and protein synthesis by Beggiatoa alba B18LD was investigated using the incorporation of radiolabeled leucine to estimate protein synthesis. Leucine was assimilated into whole cells in the presence of 6.1 mM acetate at a rate of 0.6 nmol · min^-1 · mg protein^-1, 43% of which was incorporated into the protein fraction. Protein synthesis by B. alba was unaffected by 1 mM sulfide, whether or not the cells had been preincubated with sulfide. B. alba oxidized radioactive sulfide to sulfur within 30 s of addition of the label, whether or not the organism was preinduced by sulfide. Furthermore, chloramphenicol, which inhibited protein synthesis, did not significantly inhibit sulfide oxidation by sulfide-induced or uninduced B. alba. This indicates that sulfide oxidation is a constitutive process. Enrichments of sulfur inclusions from B. alba B18LD that were analyzed by polyacrylamide gel electrophoresis demonstrated two enriched peptides with M_r values of 13,000 and 15,000. The 13,000 and 15,000 M_r peptide bands were more evident in cells grown in a medium containing sulfide than in cells from a medium lacking sulfide. Although sulfide did not increase the rate of overall protein synthesis, the synthesis of a few peptides was increased by the addition of sulfide to the growth medium. Among those, the 15,000 M_r peptide was one of the most distinctive.Non-standard abbreviations SDS-PAGE Sodium dodecyl sulfate polyacrylamide gel electrophoresis - PPO 2,5-diphenyloxazole - POPOP 1,4-bis [5-phenyl-2-oxazolyl]-benzene - BSS basal salts solution - BH Beggiatoa heterotrophic (medium) - BSO Beggiatoa sulfide oxidation (medium) - CM chloramphenicol - TCA trichloroacetic acid - M_r molecular mass     

Some characteristics of cytochromec-555 isolated from sulfide oxidizingXanthomonas sp. DY44

From a heterotrophic bacterium,Xanthomonas sp. DY44 which was previously reported to oxidize hydrogen sulfide (H₂S) to polysulfide, cytochromec-555 (cyt.c-555) responsible for oxidation of sulfide was purified by DEAE-Toyopearl and Sepadex G-75 column chromatography. Cyt.c-555 with a molecular weight of 12,500 showed maximum absorption at 555 nm (α-peak), 522 nm (β-peak) and 417 nm (γ-peak) for the reduced form which was prepared by addition of Na₂S₂O₄. Cyt.c-555 was also reduced by addition of sulfide (Na₂S and H₂S), and the oxidized products of sulfide by cyt.c-555 was identified as polysulfide. The reduced form of cyt.c-555 was suggested to be oxidized coupled with cyt.c oxidase which is tolerant to sulfide.     

Cytochromes of the green sulfur bacterium Chlorobium vibrioforme f. thiosulfatophilum. Purification,characterization and sulfur metabolism

Three cytochromes of the thiosulfate-utilizing green sulfur bacterium Chlorobium vibrioforme f. thiosulfatophilum were highly purified by ion exchange column chromatography and ammonium sulfate fractionation. All three cytochromes are located in the soluble fraction. Cytochrome c-551 (highest purity index obtained: A₂₈₀/A₄₁₆=0.39) shows maxima at 551 nm (-band), 521 nm (-band), and 416 nm (-band) for the reduced form. This cytochrome is an acidic protein with a molecular weight of 32,000, a redox potential of 150 mV, and an isoelectric point at pH 6.0. Cytochrome c-553 (highest purity index obtained: A₂₈₀/A₄₁₇=0.8) is also an acidic protein with maxima at 553,5 nm, 523,5 nm and 417 nm for the reduced form, a molecular weight of 63,000, a redox potential of 90 mV, an isoelectric point at pH 6.3, and it contains FAD as flavin component. It is autoxidizable and participates in sulfide oxidation, but cannot catalyze the reverse reaction. The cytochrome c-555 (highest purity index obtained: A₂₈₀/A₄₁₈=0.16) is a small basic protein with maxima at 555 nm, 523 nm and 418 nm (reduced form), a molecular weight of 12,500, an isoelectric point between pH 10 and 10.5, and a redox potential of 155 mV. The ratio of the cytochrome contents to each other is constant and does not change when the organism has only thiosulfate or sulfide as the main electron donor in the medium.The soluble fraction further contains the non-heme ironcontaining proteins rubredoxin and ferredoxin. The anaerobic sulfide oxidation in a growing culture of Chlorobium vibrioforme f. thiosulfatophilum is accompanied by a rapid formation of thiosulfate, which is only utilized when sulfide is no longer available, while the elemental sulfur concentration increases constantly until thiosulfate is consumed.Non-common abbreviations C Chlorobium - SDS sodium dodecylsulfate - HIPIP high-potential-iron-sulfur-protein     

Inhibition of microbiological sulfide oxidation by methanethiol and dimethyl polysulfides at natron-alkaline conditions

To avoid problems related to the discharge of sulfidic spent caustics, a biotechnological process is developed for the treatment of gases containing both hydrogen sulfide and methanethiol. The process operates at natron-alkaline conditions (>1 mol L⁻¹ of sodium- and potassium carbonates and a pH of 8.5–10) to enable the treatment of gases with a high partial CO₂ pressure. In the process, methanethiol reacts with biologically produced sulfur particles to form a complex mixture predominantly consisting of inorganic polysulfides, dimethyl disulfide (DMDS), and dimethyl trisulfide (DMTS). The effect of these organic sulfur compounds on the biological oxidation of sulfide to elemental sulfur was studied with natron-alkaliphilic bacteria belonging to the genus Thioalkalivibrio. Biological oxidation rates were reduced by 50% at 0.05 mM methanethiol, while for DMDS and DMTS, this was estimated to occur at 1.5 and 1.0 mM, respectively. The inhibiting effect of methanethiol on biological sulfide oxidation diminished due to its reaction with biologically produced sulfur particles. This reaction increases the feasibility of biotechnological treatment of gases containing both hydrogen sulfide and methanethiol at natron-alkaline conditions.     

Microbiological analysis of the population of extremely haloalkaliphilic sulfur-oxidizing bacteria dominating in lab-scale sulfide-removing bioreactors

Thiopaq biotechnology for partial sulfide oxidation to elemental sulfur is an efficient way to remove H₂S from biogases. However, its application for high-pressure natural gas desulfurization needs upgrading. Particularly, an increase in alkalinity of the scrubbing liquid is required. Therefore, the feasibility of sulfide oxidation into elemental sulfur under oxygen limitation was tested at extremely haloalkaline conditions in lab-scale bioreactors using mix sediments from hypersaline soda lakes as inoculum. The microbiological analysis, both culture dependent and independent, of the successfully operating bioreactors revealed a domination of obligately chemolithoautotrophic and extremely haloalkaliphilic sulfur-oxidizing bacteria belonging to the genus Thioalkalivibrio. Two subgroups were recognized among the isolates. The subgroup enriched from the reactors operating at pH 10 clustered with Thioalkalivibrio jannaschii–Thioalkalivibrio versutus core group of the genus Thioalkalivibrio. Another subgroup, obtained mostly with sulfide as substrate and at lower pH, belonged to the cluster of facultatively alkaliphilic Thioalkalivibrio halophilus. Overall, the results clearly indicate a large potential of the genus Thiolalkalivibrio to efficiently oxidize sulfide at extremely haloalkaline conditions, which makes it suitable for application in the natural gas desulfurization. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Nucleotide sequence accession numbers: The GenBank/EMBL accession numbers of the 16S rRNA gene sequence determined in this study are EU709849–EU709878.     

Search for polythionates in cultures of Chromatium vinosum after sulfide incubation

Cultures of Chromatium vinosum, devoid of sulfur globules, were supplemented with sulfide and incubated under anoxic conditions in the light. The concentrations of sulfide, polysulfides, thiosulfate, polythionates and elemental sulfur (sulfur rings) were monitored for 3 days by ion-chromatography and reversed-phase HPLC. While sulfide disappeared rapidly, thiosulfate and elemental sulfur (S₆, S₇ S₈ rings) were formed. After sulfide depletion, the concentration of thiosulfate decreased fairly rapidly, but elemental sulfur was oxidized very slowly to sulfate. Neither polysulfides (S _x ^2– ), polythionates (S_nO ₆ ^2– , n=4–6), nor other polysulfur compounds could be detected, which is in accordance with the fact that sulfide-grown cells were able to oxidize polysulfide without lag. The nature of the intracellular sulfur globules is discussed.     

Patterns of pyrite oxidation by different microorganisms

The bacterial-chemical oxidation of natural pyrites with different physical, chemical, and electrophysical characteristics by bacteria Acidithiobacillus ferrooxidans, Sulfobacillus thermotolerans, and the archaeon Ferroplasma acidiphilum were studied. The electrophysical characteristics of three natural pyrites differed in the K _thermoEMF value (pyrites 3, 4, hole conduction (p-type conductivity); pyrite 5, mixed type conductivity (n-p)) and in the logarithm of electric resistance. Chemical oxidation of pyrites 3 and 5 resulted in no changes of K _thermoEMF. When pyrite 4 was oxidized chemically, the K _thermoEMF values remained in the same range as in the initial sample, but the ratio of grains with different K _thermoEMF values in the sample was changed: the number of grains with a higher K _thermoEMF value increased. The same changes were also observed in the course of bacterio-chemical oxidation of pyrite 4. Of the three pyrites studied, an increase in the logarithm of resistance was observed only for chemical oxidation of pyrite 4 at 28°C. At higher experimental temperatures, the logarithm of resistance increased accordingly; more active bacterial-chemical oxidation resulted in a more pronounced increase in the logarithm of resistance than chemical oxidation. On bacterial-chemical oxidation of pyrites 3 and 5 by A. ferrooxidans and S. thermotolerans strains, iron was leached more actively than sulfur. Preferred bacterial-chemical oxidation of certain fractions from the pyrite samples was shown, namely of the pyrite 3 fraction with higher K _thermoEMF values by the F. acidiphilum strain and of a fraction from the pyrite 5 sample with medium K _thermoEMF values by the A. ferrooxidans and S. thermotolerans strains. The comparative assessment of bacterial-chemical pyrite oxidation by three types of microorganisms showed the direction of changes in the K _thermoEMF values to be the same in the case of bacteria Acidithiobacillus ferrooxidans and Sulfobacillus thermotolerans and different in the case of the archaeon Ferroplasma acidiphilum.     

Production of elemental sulfur by green and purple sulfur bacteria

The utilization of sulfide by phototrophic sulfur bacteria temporarily results in the accumulation of elemental sulfur. In the green sulfur bacteria (Chlorobiaceae), the sulfur is deposited outside the cells, whereas in the purple sulfur bacteria (Chromatiaceae) sulfur is found intracellularly. Consequently, in the latter case, sulfur is unattainable for other individuals. Attempts were made to analyze the impact of the formation of extracellular elemental sulfur compared to the deposition of intracellular sulfur.According to the theory of the continuous cultivation of microorganisms, the steady-state concentration of the limiting substrate is unaffected by the reservoir concentration (S _R).It was observed in sulfide-limited continuous cultures ofChlorobium limicola f.thiosulfatophilum that higherS _R values not only resulted in higher steady-state population densities, but also in increased steady-state concentrations of elemental sulfur. Similar phenomena were observed in sulfide-limited cultures ofChromatium vinosum.It was concluded that the elemental sulfur produced byChlorobium, althouth being deposited extracellularly, is not easily available for other individuals, and apparently remains (in part) attached to the cells. The ecological significance of the data is discussed.Non-standard abbreviations RP reducing power - BChl bacteriochlorophyll - N_cell cell material - specific growth rate - {ie52-1} maximal specific growth rate - D dilution rate - K _s saturation constant - s concentration of limiting substrate - S _R same ass but in reservoir bottle - Y yield factor - iS^o intracellular elemental sulfur - eS^o extracellular elemental sulfur - PHB poly-beta-hydroxybutyric acid     

Cytochromes,rubredoxin, and sulfur metabolism of the non-thiosulfate-utilizing green sulfur bacterium Pelodictyon luteolum

Two soluble c-type cytochromes (c-553 and c-555) and the nonheme iron-containing protein rubredoxin of the non-thiosulfate-utilizing green sulfur bacterium Pelodictyon luteolum were highly purified by ion exchange column chromatography, gel filtration and ammonium sulfate fractionation. Both cytochrome are small and basic hemoproteins, while rubredoxin is an acidic small nonheme iron protein. Cytochrome c-553 has a molecular weight of 13,000 determined by Sephacryl S-200 chromatography and of 10,700 by electrophoresis on SDS acrylamide gel, an isoelectric point at pH 10.2, a redox-potential of +220 mV. It shows maxima at 413 nm in the oxidized form, and the characteristic three maxima in the reduced state (-band at 553 nm, -band at 523 nm, -band at 417 nm). The best purity index (A ₂₈₀/A ₄₁₇) obtained was 0.18. Cytochrome c-555 (best purity index obtained: A ₂₈₀/A ₄₁₈=0.17) has an isoelectric point at pH 10.5, a molecular weight of 9,500 (by electrophoresis on SDS acrylamide gel) and a redox-potential of +160mV. The reduced form of this cytochrome shows the typical bands of c-type cytochromes at 555 (551) nm (-band), 523 nm (-band) and 418 nm (-band), while the oxidized form has the -band at 413 nm.Rubredoxin (best purity index obtained: A ₂₈₀/A ₄₉₀=3.5) is an acidic small protein. Its molecular weight estimated by gel filtration and SDS acrylamide gel electrophoresis is 27,000 and 6,300 respectively. The monomer of this protein contains one iron atom per molecule. Rubredoxin has an isoelectric point at pH 2.8 and shows maxima at 570 nm, 490 nm and 370 nm in the oxidized form.During anaerobic sulfide oxidation of a growing culture of Pelodictyon luteolum elemental sulfur is the first main product, which appears in the medium. Elemental sulfur is further oxidized to sulfate, after the available sulfide is completely consumed by the cells.Non-common abbreviations C Chlorobium - P Pelodictyon - SDS sodium dodecylsulfate - HIPIP high-potential-iron-sulfur-protein Offprint requests to: U. Fischer     

Restriction Profiles of the Chromosomal DNA from Acidithiobacillus ferrooxidans Strains Adapted to Different Oxidation Substrates

Restriction profiles of chromosomal DNA were studied in different Acidithiobacillus ferrooxidans strains grown on medium with Fe²⁺ and further adapted to another oxidation substrate (S⁰, FeS₂, or sulfide ore concentrates). The restriction endonuclease XbaI digested the chromosomal DNA from different strains into different numbers of fragments of various sizes. Adaptation of two strains (TFBk and TFN-d) to new oxidation substrates resulted in structural changes in XbaI-restriction patterns of their chromosomal DNA. Such changes in the DNA restriction patterns occurred in strain TFBk after the adaptation to precyanidated gravitational pyrite-arsenopyrite concentrate (no. 1) from the Nezhdaninskoe deposit or to copper-containing ore from the Udokanskoe deposit and also in strain TFN-d adapted to untreated pyrite-arsenopyrite concentrate (no. 2) from the Nezhdaninskoe deposit. No changes in the number or size of the XbaI-restriction patterns of chromosomal DNA were revealed in either strain TFBk cultivated on media with pyrite from the Angren and Tulun deposits or in strains TFN-d and TFO grown on media with S⁰ and pyrite. Neither were changes observed in the XbaI-restriction patterns of the DNA from strain TFV-1, isolated from the copper ore of the Volkovskoe deposit, when Fe²⁺ was substituted with alternative substrates—S⁰, pyrite or concentrate no. 2 from the ore of the Nezhdaninskoe deposit. In strain TFO, no differences in the XbaI-restriction patterns of the chromosomal DNA were revealed between the culture grown on medium containing concentrate no. 2 or the concentrate of surface-lying ore from the Olimpiadinskoe deposit and the culture grown on medium with Fe²⁺. When strain TFO was cultivated on the ore concentrate from deeper horizons of the Olimpiadinskoe deposit, which are characterized by lower oxidation degrees and high antimony content, mutant TFO-2 differing from the parent strain in the chromosomal DNA structure was isolated. The correlation between the lability of the chromosomal DNA structure in A. ferrooxidans strains and the physical and chemical peculiarities of the isolation substrate and habitat is discussed.     

Microbial desulfurization of coal and oxidation of pure pyrite byThiobacillus ferrooxidans andAcidianus brierleyi

Summary Thiobacillus ferrooxidans andAcidianus brierleyi were capable of oxidizing pure pyrite as well as oxidizing sulfur in coal. First order reactions were assumed in the kinetic analysis performed. For oxidation of pure pyrite the rate constant was higher forA. brierleyi than forT. ferrooxidans. For sulfur removal from coal the values of the rate constants were comparable for the two microorganisms.