The role of Hox hydrogenase in the H2 metabolism of Thiocapsa roseopersicina

The purple sulfur phototrophic bacterium Thiocapsa roseopersicina BBS synthesizes at least three NiFe hydrogenases (Hox, Hup, Hyn). We characterized the physiological H₂ consumption/evolution reactions in mutants having deletions of the structural genes of two hydrogenases in various combinations. This made possible the separation of the functionally distinct roles of the three hydrogenases. Data showed that Hox hydrogenase (unlike the Hup and Hyn hydrogenases) catalyzed the dark fermentative H₂ evolution and the light-dependent H₂ production in the presence of thiosulfate. Both Hox⁺ and Hup⁺ mutants demonstrated light-dependent H₂ uptake stimulated by CO₂ but only the Hup⁺ mutant was able to mediate O₂-dependent H₂ consumption in the dark. The ability of the Hox⁺ mutant to evolve or consume hydrogen was found to depend on a number of interplaying factors including both growth and reaction conditions (availability of glucose, sulfur compounds, CO₂, H₂, light). The study of the redox properties of Hox hydrogenase supported the reversibility of its action. Based on the results a scheme is suggested to describe the role of Hox hydrogenase in light-dependent and dark hydrogen metabolism in T. roseopersicina BBS.     

The role of Hox hydrogenase in the H2 metabolism of Thiocapsa roseopersicina

The purple sulfur phototrophic bacterium Thiocapsa roseopersicina BBS synthesizes at least three NiFe hydrogenases (Hox, Hup, Hyn). We characterized the physiological H(2) consumption/evolution reactions in mutants having deletions of the structural genes of two hydrogenases in various combinations. This made possible the separation of the functionally distinct roles of the three hydrogenases. Data showed that Hox hydrogenase (unlike the Hup and Hyn hydrogenases) catalyzed the dark fermentative H(2) evolution and the light-dependent H(2) production in the presence of thiosulfate. Both Hox(+) and Hup(+) mutants demonstrated light-dependent H(2) uptake stimulated by CO(2) but only the Hup(+) mutant was able to mediate O(2)-dependent H(2) consumption in the dark. The ability of the Hox(+) mutant to evolve or consume hydrogen was found to depend on a number of interplaying factors including both growth and reaction conditions (availability of glucose, sulfur compounds, CO(2), H(2), light). The study of the redox properties of Hox hydrogenase supported the reversibility of its action. Based on the results a scheme is suggested to describe the role of Hox hydrogenase in light-dependent and dark hydrogen metabolism in T. roseopersicina BBS.     

Growth of thermophilic, obligatorily chemolithoautotrophic hydrogen-oxidizing bacteria related to Hydrogenobacter with thiosulfate and elemental sulfur as electron and energy source

Abstract Strains related to Hydrogenobacter , a genus of thermophilic, obligatorily chemolithoautotrophic bacteria, were able to utilize elemental sulfur or thiosulfate, as well as molecular hydrogen, as sole electron and energy source. Extracellular elemental sulfur was produced as an intermediate during oxidation of thiosulfate. Growth with thiosulfate alone was strongly microaerophilic, whereas no hydrogenase activity was detected. Mixolithotrophic growth with both hydrogen and thiosulfate was faster than with hydrogen alone, and the cells harbored a hydrogenase activity comparable to that of cells grown under hydrogen without thiosulfate.     

The HydS C-terminal domain of the Thiocapsa bogorovii HydSL hydrogenase is involved in membrane anchoring and electron transfer

Thiocapsa bogorovii BBS (former name Thiocapsa roseopersicina) contains HydSL hydrogenase belonging to 1e subgroup of NiFe hydrogenases (isp-type). The operon of these hydrogenases contains gene for small subunit (hydS), gene for large subunit (hupL), and genes isp1 and isp2 between them. It is predicted that last two genes code electron transport careers for electron transfer from/to HydSL hydrogenase. However, the interaction between them is unclear. The aim of this study was to determine structural and functional role of T. bogorovii HydS C-terminal end. For this purpose, we modelled all subunits of the complex HydS-HydL-Isp1-Isp2. Hydrophobicity surface analysis of the Isp1 model revealed highly hydrophobic helices suggesting potential membrane localization, as well as the hydrophilic C-terminus, which is likely localized outside of membrane. Isp1 model was docked with models of full length and C-terminal truncated HydSL hydrogenases and results illustrate the possibility of HydSL membrane anchoring via transmembrane Isp1 with essential participation of C-terminal end of HydS in the interaction. C-terminal end of HydS subunit was deleted and our studies revealed that the truncated HydSL hydrogenase detached from cellular membranes in contrast to native hydrogenase. It is known that HydSL hydrogenase in T. bogorovii performs the reaction of elemental sulfur reduction (S₀ + H₂ = ≥H₂S). Cells with truncated HydS produced much less H₂S in the presence of H₂ and S₀. Thus, our data support the conclusion that C-terminal end of HydS subunit participates in interaction of HydSL hydrogenase with Isp1 protein for membrane anchoring and electron transfer.     

Control of Hydrogen Photoproduction by the Proton Gradient Generated by Cyclic Electron Flow in Chlamydomonas reinhardtii

Hydrogen photoproduction by eukaryotic microalgae results from a connection between the photosynthetic electron transport chain and a plastidial hydrogenase. Algal H₂ production is a transitory phenomenon under most natural conditions, often viewed as a safety valve protecting the photosynthetic electron transport chain from overreduction. From the colony screening of an insertion mutant library of the unicellular green alga Chlamydomonas reinhardtii based on the analysis of dark-light chlorophyll fluorescence transients, we isolated a mutant impaired in cyclic electron flow around photosystem I (CEF) due to a defect in the Proton Gradient Regulation Like1 (PGRL1) protein. Under aerobiosis, nonphotochemical quenching of fluorescence (NPQ) is strongly decreased in pgrl1. Under anaerobiosis, H₂ photoproduction is strongly enhanced in the pgrl1 mutant, both during short-term and long-term measurements (in conditions of sulfur deprivation). Based on the light dependence of NPQ and hydrogen production, as well as on the enhanced hydrogen production observed in the wild-type strain in the presence of the uncoupling agent carbonyl cyanide p-trifluoromethoxyphenylhydrazone, we conclude that the proton gradient generated by CEF provokes a strong inhibition of electron supply to the hydrogenase in the wild-type strain, which is released in the pgrl1 mutant. Regulation of the trans-thylakoidal proton gradient by monitoring pgrl1 expression opens new perspectives toward reprogramming the cellular metabolism of microalgae for enhanced H₂ production.     

Rational redesign of the ferredoxin-NADP+-oxido-reductase/ferredoxin-interaction for photosynthesis-dependent H2-production

Utilization of electrons from the photosynthetic water splitting reaction for the generation of biofuels, commodities as well as application in biotransformations requires a partial rerouting of the photosynthetic electron transport chain. Due to its rather negative redox potential and its bifurcational function, ferredoxin at the acceptor side of Photosystem 1 is one of the focal points for such an engineering. With hydrogen production as model system, we show here the impact and potential of redox partner design involving ferredoxin (Fd), ferredoxin-oxido-reductase (FNR) and [FeFe]?hydrogenase HydA1 on electron transport in a future cyanobacterial design cell of Synechocystis PCC 6803. X-ray-structure-based rational design and the allocation of specific interaction residues by NMR-analysis led to the construction of Fd- and FNR-mutants, which in appropriate combination enabled an about 18-fold enhanced electron flow from Fd to HydA1 (in competition with equimolar amounts of FNR) in in vitro assays. The negative impact of these mutations on the Fd-FNR electron transport which indirectly facilitates H₂ production (with a contribution of ≤42% by FNR variants and ≤23% by Fd-variants) and the direct positive impact on the Fd-HydA1 electron transport (≤23% by Fd-mutants) provide an excellent basis for the construction of a hydrogen-producing design cell and the study of photosynthetic efficiency-optimization with cyanobacteria.     

Dissimilatory Oxidation and Reduction of Elemental Sulfur in Thermophilic Archaea

The oxidation and reduction of elemental sulfur and reduced inorganic sulfur species are some of the most important energy-yielding reactions for microorganisms living in volcanic hot springs, solfataras, and submarine hydrothermal vents, including both heterotrophic, mixotrophic, and chemolithoautotrophic, carbon dioxide-fixing species. Elemental sulfur is the electron donor in aerobic archaea like Acidianus and Sulfolobus. It is oxidized via sulfite and thiosulfate in a pathway involving both soluble and membrane-bound enzymes. This pathway was recently found to be coupled to the aerobic respiratory chain, eliciting a link between sulfur oxidation and oxygen reduction at the level of the respiratory heme copper oxidase. In contrast, elemental sulfur is the electron acceptor in a short electron transport chain consisting of a membrane-bound hydrogenase and a sulfur reductase in (facultatively) anaerobic chemolithotrophic archaea Acidianus and Pyrodictium species. It is also the electron acceptor in organoheterotrophic anaerobic species like Pyrococcus and Thermococcus, however, an electron transport chain has not been described as yet. The current knowledge on the composition and properties of the aerobic and anaerobic pathways of dissimilatory elemental sulfur metabolism in thermophilic archaea is summarized in this contribution.     

Properties of the solubilized membrane-bound hydrogenase from the photosynthetic bacterium Rhodospirillum rubrum.

The membrane-bound hydrogenase of the photosynthetic bacterium Rhodospirillum rubrum has been purified 490-fold with a yield of 5.8%. The enzyme was homogeneous by disc gel electrophoresis. A method for the permanent, oxygen-insensitive, staining of hydrogenase on polyacrylamide gels is described. The enzyme is a monomer of molecular weight about 66,000 containing four iron and four acid-labile sulfur atoms per molecule. The electron paramagnetic resonance spectrum at 20 °K exhibits a strong signal in the oxidized state only with g > 2—this is characteristic of high potential iron-sulfur protein. The hydrogenase is thermostable and also resistant to both denaturation agents and oxygen inactivation. Carbon monoxide reversibly inhibits the enzyme but metal-complexing and thiol-blocking reagents have little effect on activity. The enzyme will catalyze both H₂ evolution and H₂ uptake in the presence of many artificial electron carriers but the two activities differ in their pH optima. There is a correlation between H₂ evolution activity and the redox potential of the mediator dye. Ferredoxins and pyridine nucleotides do not readily interact with the hydrogenase. We have shown that irradiation of a solution containing methyl viologen, EDTA, proflavin, and R. rubrum hydrogenase will evolve hydrogen continuously for over 9 h. However, the enzyme evolves hydrogen at only very low rates from in vitro chloroplast-ferredoxin and chloroplast-methyl viologen systems. R. rubrum hydrogenase has a number of properties in common with the hydrogenases purified from two other photosynthetic bacteria, Chromatium and Thiocapsa, but is distinct from the hydrogenases of nonphotosynthetic bacteria.     

The role of PGR5 in the redox poising of photosynthetic electron transport

The pgr5 mutant of Arabidopsis thaliana has been described as being deficient in cyclic electron flow around photosystem I, however, the precise role of the PGR5 protein remains unknown. To address this issue, photosynthetic electron transport was examined in intact leaves of pgr5 and wild type A. thaliana. Based on measurements of the kinetics of P700 oxidation in far red light and re-reduction following oxidation in the presence of DCMU, we conclude that this mutant is able to perform cyclic electron flow at a rate similar to the wild type. The PGR5 protein is therefore not essential for cyclic flow. However, cyclic flow is affected by the pgr5 mutation under conditions where this process is normally enhanced in wild type leaves, i.e. high light or low CO₂ concentrations resulted in enhancement of cyclic electron flow. This suggests a different capacity to regulate cyclic flow in response to environmental stimuli in the mutant. We also show that the pgr5 mutant is affected in the redox poising of the chloroplast, with the electron transport chain being substantially reduced under most conditions. This may result in defective feedback regulation of photosynthetic electron transport under some conditions, thus providing a rationale for the reduced efficiency of cyclic electron flow.     

On the energetics of the photosyntheses in green sulfur bacteria

The quantum efficiency of photosynthesis by the green sulfur bacterium, Chlorobium thiosulfatophilum, has been determined in systems in which thiosulfate, tetrathionate, and molecular hydrogen served as electron donors. It was found that about 10 ± 1 quanta are used for the assimilation of 1 molecule of CO₂, and that the quantum number is independent of the nature of the electron donor. These results are considered as support for the view that also in the bacterial photosyntheses the primary photochemical reaction consists in the photolysis of H₂O, and that the chemical energy released during the oxidation of the electron donor is not utilized for CO₂ assimilation. Hence the photosynthetic processes of the green sulfur bacteria are thermodynamically less efficient than is green plant photosynthesis.     

Capacity of Azospirillum thiophilum for lithotrophic growth coupled to oxidation of reduced sulfur compounds

Capacity for lithotrophic growth coupled to oxidation of reduced sulfur compounds was revealed in an Azospirillum strain, A. thiophilum BV-S ^T . Oxygen concentration in the medium was the major factor determining the type of energy metabolism (organotrophic or lithotrophic) in the presence of thiosulfate. Under aerobic conditions, metabolism of A. thiophilum BV-S^T was organoheterotrophic, with thiosulfate oxidation to tetrathionate resulting from the interaction with reactive oxygen species, mostly H₂O₂, which was formed in the electron transport chain in the course of oxidation of organic electron donors. Under microaerobic conditions (2 mg/L O₂ in liquid medium), A. thiophilum BV-S^T carried out lithoheterotrophic (mixotrophic) metabolism; enzymes of the dissimilatory type of sulfur metabolism were responsible for thiosulfate oxidation to tetrathionate and sulfate. Two enzyme systems were found in the cells: thiosulfate dehydrogenase, which catalyzes incomplete oxidation of thiosulfate to tetrathionate and the thiosulfate-oxidizing Sox enzyme complex, which is involved in complete oxidation of thiosulfate to sulfate. The genetic determinant of a Sox complex component in A. thiophilum BV-S^T was revealed. The soxB gene was found, and its expression under microaerobic conditions was observed to increase 32-fold compared to aerobic cultivation.     

A new ecological adaptation to high sulfide by a Hydrogenobacter sp. growing on sulfur compounds but not on hydrogen

Thermophilic bacteria were isolated from a sulfide-rich, neutral hot spring in Iceland on gelrite minimal medium with 16 mM thiosulfate. The isolates were aerobic, obligate chemolithoautotrophs and used thiosulfate and sulfur as electron donors, producing sulfate from both substrates. No growth was observed with hydrogen as the sole electron donor, and no hydrogenase activity was detected. The cells were gram-negative and usually single, 4—5 μm long and 0.7 μm in diameter and formed sulfur globules after a few days of incubation. By SSU rRNA sequence comparisons, the bacterium was placed in the genus Hydrogenobacter with the closest relative to be Calderobacterium hydrogenophilum with 98.3% sequence similarity. This novel bacterium shows an ecological adaptation to high sulfide springs and is differentiated from its closest known relatives by lack of H₂ oxidation, deposition of sulfur and lower growth temperature.     

A Second Soluble Hox-Type NiFe Enzyme Completes the Hydrogenase Set in Thiocapsa roseopersicina BBS

Three functional NiFe hydrogenases were previously characterized in Thiocapsa roseopersicina BBS: two of them are attached to the periplasmic membrane (HynSL and HupSL), and one is localized in the cytoplasm (HoxEFUYH). The ongoing genome sequencing project revealed the presence of genes coding for another soluble Hox-type hydrogenase enzyme (hox2FUYH). Hox2 is a heterotetrameric enzyme; no indication for an additional subunit was found. Detailed comparative in vivo and in vitro activity and expression analyses of HoxEFUYH (Hox1) and the newly discovered Hox2 enzyme were performed. Functional differences between the two soluble NiFe hydrogenases were disclosed. Hox1 seems to be connected to both sulfur metabolism and dark/photofermentative processes. The bidirectional Hox2 hydrogenase was shown to be metabolically active under specific conditions: it can evolve hydrogen in the presence of glucose at low sodium thiosulfate concentration. However, under nitrogen-fixing conditions, it can oxidize H₂ but less than the other hydrogenases in the cell.Hydrogenases are metalloenzymes involved in microbial hydrogen metabolism. A great variety of them have been identified and studied in various microorganisms and grouped on the basis of their metal content as NiFe, FeFe, and iron-sulfur cluster free hydrogenases (10, 42, 43). The basic protein structure of NiFe hydrogenases is heterodimeric, while FeFe hydrogenases are mostly composed of a single amino acid chain with multiple iron-sulfur clusters (28, 43, 44). Well-defined maturation proteins assist for the assembly and activation of hydrogenase enzymes; NiFe hydrogenases require a more complex accessory machinery than FeFe enzymes (2, 3, 24).Thiocapsa roseopersicina BBS is a photosynthetic purple sulfur bacterium belonging to the Chromatiaceae family (4). It prefers to utilize reduced sulfur compounds for anaerobic photochemolithoautotrophic growth, but simple organic substrates such as glucose or acetate can be also used as extra carbon, energy, and electron sources. It can be cultivated under aerobic (nonphotosynthetic) conditions in the presence of organic compounds. In the absence of other nitrogen sources, it is able to fix molecular nitrogen; this process is accompanied by H₂ production. T. roseopersicina was earlier shown to possess at least three NiFe hydrogenases varying in their in vivo functions, localizations, and compositions. Hyn and Hup hydrogenases are attached to the membrane facing the periplasmic side (6, 18, 30). Hyn is a bidirectional enzyme with extraordinary stability (17). Recent study has demonstrated that the HynSL subunits are physiologically connected to cellular redox processes via the Isp1 and Isp2 proteins, which play an essential role in electron transfer (27). The second membrane-associated enzyme, Hup, is involved in H₂ oxidation and shows homology to uptake hydrogenases, which recycle H₂ produced by the nitrogenase enzyme complex or present in the environment. Next to the hydrogenase small and large subunits (HupSL), a b-type cytochrome, HupC, was demonstrated to be part of the in vivo active enzyme as a transmitter of electrons to the quinone pool (27). In several bacteria, e.g., Rhodobacter capsulatus (7) and Ralstonia eutropha (15, 20), the expression of the hydrogenase(s) was shown to be regulated by the hydrogen level in the environment. The genes encoding the hydrogen-sensing system also exist in T. roseopersicina (hupUV, hupT, and hupR), but the hupTUV genes proved to be silent in the wild-type strain—only hupR is expressed—which is why expression of hupSL genes is constitutive (16).A Hox-type soluble hydrogenase was also identified in T. roseopersicina (31); it is a representative of the bidirectional heteromultimeric cytoplasmic NiFe hydrogenases (37, 39). Enzymes belonging to this group are basically composed of two moieties: hydrogenase (HoxYH) and diaphorase (HoxFU) heterodimers. Additional subunits were identified in few cases. In R. eutropha H16, two HoxI proteins completing the Hox complex were suggested to provide a binding domain for NADPH (5). HoxE has been identified as the fifth subunit of heteropentameric NAD⁺-reducing Hox hydrogenases in several cyanobacteria, Allochromatium vinosum and T. roseopersicina (21, 31, 37). In-frame deletion of the hoxE gene ceased both the H₂-producing and -oxidizing activities of Hox in vivo, but these were not affected in vitro. Consequently, an electron transfer role of the HoxE subunit was suggested (31, 32).The possibility of the presence of further hydrogenases in T. roseopersicina was noted few years ago (31). In the hynSL hupSL hoxH triple-mutant strain ({"type":"entrez-nucleotide","attrs":{"text":"GB112131","term_id":"336599610","term_text":"GB112131"}}GB112131), a small in vivo and in vitro hydrogenase activity could be measured under photomixotrophic growth conditions (both CO₂ and organic compounds are used for growth) at the late growth phase. This residual activity could not be detected in the hypF mutant strain (M539). Since HypF protein has an essential role in the maturation process of all NiFe hydrogenases (9), these results suggested the presence of a previously unknown hydrogenase. Here we describe the identification and characterization of the second Hox-type hydrogenase, emphasizing the functional similarities and differences between the two soluble enzymes of this bacterium. In order to distinguish between the two Hox-type enzymes unequivocally, the HoxEFUYH complex will be renamed Hox1 and the newly described Hox2FUYH enzyme is called Hox2.     

The effect of sulfur compounds on H<Subscript>2</Subscript> evolution/consumption reactions,mediated by various hydrogenases,in the purple sulfur bacterium, <Emphasis Type="Italic">Thiocapsa roseopersicina</Emphasis>

The influence of reduced sulfur compounds (including stored S⁰) on H₂ evolution/consumption reactions in the purple sulfur bacterium, Thiocapsa roseopersicina BBS, was studied using mutants containing only one of the three known [NiFe] hydrogenase enzymes: Hox, Hup or Hyn. The observed effects depended on the kind of hydrogenase involved. The mutant harbouring Hox hydrogenase was able to use S₂O₃²⁻, SO₃²⁻, S²⁻ and S⁰ as electron donors for light-dependent H₂ production. Dark H₂ evolution from organic substrates via Hox hydrogenase was inhibited by S⁰. Under light conditions, endogenous H₂ uptake by Hox or Hup hydrogenases was suppressed by S compounds. СО₂-dependent H₂ uptake by Hox hydrogenase in the light required the additional presence of S compounds, unlike the Hup-mediated process. Dark H₂ consumption via Hyn hydrogenase was connected to utilization of S⁰ as an electron acceptor and resulted in the accumulation of H₂S. In wild type BBS, with high levels of stored S⁰, dark H₂ production from organic substrates was significantly lower, but H₂S accumulation significantly higher, than in the mutant GB1121(Hox⁺). There is a possibility that H₂ produced via Hox hydrogenase is consumed by Hyn hydrogenase to reduce S⁰.     

The Role of Hydrogen for Sulfurimonas denitrificans’ Metabolism

Sulfurimonas denitrificans was originally isolated from coastal marine sediments. It can grow with thiosulfate and nitrate or sulfide and oxygen. Recently sequencing of its genome revealed that it encodes periplasmic and cytoplasmic [NiFe]-hydrogenases but the role of hydrogen for its metabolism has remained unknown. We show the first experimental evidence that S. denitrificans can indeed express a functional hydrogen uptake active hydrogenase and can grow on hydrogen. In fact, under the provided conditions it grew faster and denser on hydrogen than on thiosulfate alone and even grew with hydrogen in the absence of reduced sulfur compounds. In our experiments, at the time points tested, the hydrogen uptake activity appeared to be related to the periplasmic hydrogenase and not to the cytoplasmic hydrogenase. Our data suggest that under the provided conditions S. denitrificans can grow more efficiently with hydrogen than with thiosulfate.     

Generation of a proton gradient in Desulfovibrio vulgaris

Washed cells of Desulfovibrio vulgaris strain Marburg oxidized H₂, formate, lactate or pyruvate with sulfate, sulfite, trithionate, thiosulfate or oxygen as electron acceptor. CuCl₂ as an inhibitor of periplasmic hydrogenase inhibited H₂ and formate oxidation with sulfur compounds, and lactate oxidation in H₂-grown, but not in lactate-grown cells. H₂ oxidation was sensitive to O₂ concentrations above 2% saturation. Carbon monoxide inhibited the oxidation of all substrates tested. Additions of micromolar H₂ pulses to cells incubated in KCl in the presence of various sulfur compounds (reductant pulse method) resulted in a reversible acidification. This proton release was stimulated by thiocyanate, methyl triphenylphosphonium (MTPP⁺) or valinomycin plus EDTA, and completely inhibited by the uncoupler carbonylcyanide m-chlorophenylhydrazone (CCCP), CuCl₂ or carbon monoxide. The extrapolated H⁺/H₂ ratios obtained with sulfate, sulfite, trithionate or thiosulfate varied from 1.0 to 1.7. Micromolar additions of O₂ to cells incubated in the presence of excess of electron donor (oxidant pulse method) caused proton translocation with extrapolated H⁺/H₂ ratios of 3.9 with H₂, 1.6 with lactate and 2.4 with pyruvate. Since a periplasmic hydrogenase can release at maximum 2 H⁺/H₂, it is concluded that D. vulgaris is able to generate a proton gradient by vectorial proton translocation across the cytoplasmic membrane and by extracellular proton release by a periplasmic hydrogenase.     

A membrane-bound multienzyme, hydrogen-oxidizing, and sulfur-reducing complex from the hyperthermophilic bacterium Aquifex aeolicus

Aquifex aeolicus is a hyperthermophilic, chemolithoautotrophic, hydrogen-oxidizing, and microaerophilic bacterium growing at 85 degrees C. We have shown that it can grow on an H2/S degrees medium and produce H2S from sulfur in the later exponential phase. The complex carrying the sulfur reducing activity (electron transport from H2 to S degrees ) has been purified and characterized. It is a membrane-bound multiprotein complex containing a [NiFe] hydrogenase and a sulfur reductase connected via quinones. The sulfur reductase is encoded by an operon annotated dms (dimethyl sulfoxide reductase) that we have renamed sre and is composed of three subunits. Sequence analysis showed that it belongs to the Me2SO reductase molybdoenzyme family and is similar to the sulfur/polysulfide/thiosulfate/tetrathionate reductases. The study of catalytic properties clearly demonstrated that it can reduce tetrathionate, sulfur, and polysulfide, but cannot reduce Me2SO and thiosulfate, and that NADPH increases the sulfur reducing activity. To date, this is the first characterization of a supercomplex from a bacterium that couples hydrogen oxidation and sulfur reduction. The distinctive feature in A. aeolicus is the cytoplasmic localization of the sulfur reduction, which is in accordance with the presence of sulfur globules in the cytoplasm. Association of this sulfur-reducing complex with a hydrogen-oxygen pathway complex (hydrogenase I, bc1 complex) in the membrane suggests that subcomplexes involved in respiratory chains in this bacterium are part of supramolecular organization.     

Acclimation of green algae to sulfur deficiency: underlying mechanisms and application for hydrogen production

Hydrogen is definitely one of the most acceptable fuels in the future. Some photosynthetic microorganisms, such as green algae and cyanobacteria, can produce hydrogen gas from water by using solar energy. In green algae, hydrogen evolution is coupled to the photosynthetic electron transport in thylakoid membranes via reaction catalyzed by the specific enzyme, (FeFe)-hydrogenase. However, this enzyme is highly sensitive to oxygen and can be quickly inhibited when water splitting is active. A problem of incompatibility between the water splitting and hydrogenase reaction can be overcome by depletion of algal cells of sulfur which is essential element for life. In this review the mechanisms underlying sustained hydrogen photoproduction in sulfur deprived C. reinhardtii and the recent achievements in studying of this process are discussed. The attention is focused on the biophysical and physiological aspects of photosynthetic response to sulfur deficiency in green algae.     

A new ecological adaptation to high sulfide by a Hydrogenobacter sp. growing on sulfur compounds but not on hydrogen

Thermophilic bacteria were isolated from a sulfide-rich, neutral hot spring in Iceland on gelrite minimal medium with 16 mM thiosulfate. The isolates were aerobic, obligate chemolithoautotrophs and used thiosulfate and sulfur as electron donors, producing sulfate from both substrates. No growth was observed with hydrogen as the sole electron donor, and no hydrogenase activity was detected. The cells were gram-negative and usually single, 4-5 microm long and 0.7 microm in diameter and formed sulfur globules after a few days of incubation. By SSU rRNA sequence comparisons, the bacterium was placed in the genus Hydrogenobacter with the closest relative to be Calderobacterium hydrogenophilum with 98.3% sequence similarity. This novel bacterium shows an ecological adaptation to high sulfide springs and is differentiated from its closest known relatives by lack of H2 oxidation, deposition of sulfur and lower growth temperature.