Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 19 de 19
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Bioelectrochemistry ; 145: 108072, 2022 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-35144167

RESUMO

Utilization of hyperthermophilic electro-active microorganisms in microbial electrolysis cells (MECs) that are used for hydrogen production from organic wastes offers significant advantages, such as increased reaction rate and enhanced degradation of insoluble materials. However, only a limited number of hyperthermophilic bioelectrochemical systems have been investigated so far. This study is the first to illustrate hydrogen production in hyperthermophilic MECs with a maximum rate of 0.57 ±â€¯0.06 m3 H2/m3d, where an iron reducing archaeon, Geoglobus acetivorans, was used as inoculum. In fact, this is the first study to report that G. acetivorans, as the fourth hyperthermophilic electro-active archaeon. In single chamber MECs operated at 80 °C with a set potential of 0.7 V, a peak current density of 1.53 ±â€¯0.24 A/m2 has been attained and this is the highest record of current produced by pure culture hyperthermophilic microorganisms. Turnover cyclic voltammetry curve illustrated a sigmoidal shape (midpoint of -0.40 V vs. Ag/AgCl), and together with linear relation of scan rate and peak anodic current, proves the biofilm attachment to the anode and its capability of direct electron transfer. Along with simple substrate (acetate), G. acetivorans effectively utilized dark fermentation effluent for hydrogen production in MECs.


Assuntos
Archaeoglobales , Fontes de Energia Bioelétrica , Archaea/metabolismo , Eletrodos , Eletrólise , Elétrons , Hidrogênio/metabolismo
2.
Mol Microbiol ; 114(1): 17-30, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32080908

RESUMO

Class I benzoyl-CoA reductases (BCRs) are oxygen-sensitive key enzymes in the degradation of monocyclic aromatic compounds in anaerobic prokaryotes. They catalyze the ATP-dependent reductive dearomatization of their substrate to cyclohexa-1,5-diene-1-carboxyl-CoA (1,5-dienoyl-CoA). An aromatizing 1,5-dienoyl-CoA oxidase (DCO) activity has been proposed to protect BCRs from oxidative damage, however, the gene and its product involved have not been identified, yet. Here, we heterologously produced a DCO from the hyperthermophilic euryarchaeon Ferroglobus placidus that coupled the oxidation of two 1,5-dienoyl-CoA to benzoyl-CoA to the reduction of O2 to water at 80°C. DCO showed similarities to members of the old yellow enzyme family and contained FMN, FAD and an FeS cluster as cofactors. The O2 -dependent activation of inactive, reduced DCO is assigned to a redox thiol switch at Eo ' = -3 mV. We propose a catalytic cycle in which the active site FMN/disulfide redox centers are reduced by two 1,5-dienoyl-CoA (reductive half-cycle), followed by two consecutive two-electron transfer steps to molecular oxygen via peroxy- and hydroxyflavin intermediates yielding water (oxidative half-cycle). This work identified the enzyme involved in a unique oxygen detoxification process for an oxygen-sensitive catabolic enzyme.


Assuntos
Archaeoglobales/metabolismo , Metabolismo Energético/fisiologia , Hidroliases/metabolismo , Hidrocarbonetos Aromáticos/metabolismo , Oxigênio/metabolismo , Archaeoglobales/enzimologia , Archaeoglobales/genética , Domínio Catalítico/fisiologia , Dissulfetos/metabolismo , Flavinas/metabolismo , Hidroliases/genética , Hidrólise , Oxirredução
3.
Extremophiles ; 22(6): 877-888, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30062607

RESUMO

A novel type 1 geranylgeranyl pyrophosphate synthase GACE1337 has been identified within the genome of a newly identified hyperthermophilic archaeon Geoglobus acetivorans. The enzyme has been cloned and over-expressed in Escherichia coli. The recombinant enzyme has been biochemically and structurally characterized. It is able to catalyze the synthesis of geranylgeranyl pyrophosphate as a major product and of farnesyl pyrophosphate in smaller amounts, as measured by gas chromatography-mass spectrometry at an elevated temperature of 60 °C. Its ability to produce two products is consistent with the fact that GACE1337 is the only short-chain isoprenyl diphosphate synthase in G. acetivorans. Attempts to crystallize the enzyme were successful only at 37 °C. The three-dimensional structure of GACE1337 was determined by X-ray diffraction to 2.5 Å resolution. A comparison of its structure with those of related enzymes revealed that the Geoglobus enzyme has the features of both type I and type III geranylgeranyl pyrophosphate synthases, which allow it to regulate the product length. The active enzyme is a dimer and has three aromatic amino acids, two Phe, and a Tyr, located in the hydrophobic cleft between the two subunits. It is proposed that these bulky residues play a major role in the synthetic reaction by controlling the product elongation.


Assuntos
Proteínas Arqueais/química , Archaeoglobales/enzimologia , Dimetilaliltranstransferase/química , Proteínas Arqueais/genética , Proteínas Arqueais/metabolismo , Dimetilaliltranstransferase/genética , Dimetilaliltranstransferase/metabolismo , Domínios Proteicos
4.
Bioelectrochemistry ; 119: 142-149, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-28992595

RESUMO

Few microorganisms have been examined for current generation under thermophilic (40-65°C) or hyperthermophilic temperatures (≥80°C) in microbial electrochemical systems. Two iron-reducing archaea from the family Archaeoglobaceae, Ferroglobus placidus and Geoglobus ahangari, showed electro-active behavior leading to current generation at hyperthermophilic temperatures in single-chamber microbial electrolysis cells (MECs). A current density (j) of 0.68±0.11A/m2 was attained in F. placidus MECs at 85°C, and 0.57±0.10A/m2 in G. ahangari MECs at 80°C, with an applied voltage of 0.7V. Cyclic voltammetry (CV) showed that both strains produced a sigmoidal catalytic wave, with a mid-point potential of -0.39V (vs. Ag/AgCl) for F. placidus and -0.37V for G. ahangari. The comparison of CVs using spent medium and turnover CVs, coupled with the detection of peaks at the same potentials in both turnover and non-turnover conditions, suggested that mediators were not used for electron transfer and that both archaea produced current through direct contact with the electrode. These two archaeal species, and other hyperthermophilic exoelectrogens, have the potential to broaden the applications of microbial electrochemical technologies for producing biofuels and other bioelectrochemical products under extreme environmental conditions.


Assuntos
Archaeoglobales/química , Condutividade Elétrica , Eletrólise , Temperatura , Archaeoglobales/fisiologia , Biofilmes
5.
Appl Environ Microbiol ; 81(8): 2735-44, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25662973

RESUMO

The hyperthermophilic archaeon Ferroglobus placidus can utilize a wide variety of electron donors, including hydrocarbons and aromatic compounds, with Fe(III) serving as an electron acceptor. In Fe(III)-reducing bacteria that have been studied to date, this process is mediated by c-type cytochromes and type IV pili. However, there currently is little information available about how this process is accomplished in archaea. In silico analysis of the F. placidus genome revealed the presence of 30 genes coding for putative c-type cytochrome proteins (more than any other archaeon that has been sequenced to date), five of which contained 10 or more heme-binding motifs. When cell extracts were analyzed by SDS-PAGE followed by heme staining, multiple bands corresponding to c-type cytochromes were detected. Different protein expression patterns were observed in F. placidus cells grown on soluble and insoluble iron forms. In order to explore this result further, transcriptomic studies were performed. Eight genes corresponding to multiheme c-type cytochromes were upregulated when F. placidus was grown with insoluble Fe(III) oxide compared to soluble Fe(III) citrate as an electron acceptor. Numerous archaella (archaeal flagella) also were observed on Fe(III)-grown cells, and genes coding for two type IV pilin-like domain proteins were differentially expressed in Fe(III) oxide-grown cells. This study provides insight into the mechanisms for dissimilatory Fe(III) respiration by hyperthermophilic archaea.


Assuntos
Proteínas Arqueais/genética , Archaeoglobales/genética , Citocromos c/genética , Compostos Férricos/metabolismo , Regulação da Expressão Gênica , Genoma Arqueal , Proteínas Arqueais/metabolismo , Archaeoglobales/metabolismo , Citocromos c/metabolismo , Dados de Sequência Molecular , Oxirredução , Proteoma , Análise de Sequência de DNA , Transcriptoma
6.
Environ Microbiol ; 17(9): 3289-300, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25630364

RESUMO

The Fe(III)-respiring Ferroglobus placidus is the only known archaeon and hyperthermophile for which a complete degradation of aromatic substrates to CO2 has been reported. Recent genome and transcriptome analyses proposed a benzoyl-coenzyme A (CoA) degradation pathway similar to that found in the phototrophic Rhodopseudomonas palustris, which involves a cyclohex-1-ene-1-carboxyl-CoA (1-enoyl-CoA) forming, ATP-dependent key enzyme benzoyl-CoA reductase (BCR). In this work, we demonstrate, by first in vitro studies, that benzoyl-CoA is ATP-dependently reduced by two electrons to cyclohexa-1,5-dienoyl-CoA (1,5-dienoyl-CoA), which is further degraded by hydration to 6-hydroxycyclohex-1-ene-1-carboxyl-CoA (6-OH-1-enoyl-CoA); upon addition of NAD(+) , the latter was subsequently converted to ß-oxidation intermediates. The four candidate genes of BCR were heterologously expressed, and the enriched, oxygen-sensitive enzyme catalysed the two-electron reduction of benzoyl-CoA to 1,5-dienoyl-CoA. A gene previously assigned to a 2,3-didehydropimeloyl-CoA hydratase was heterologously expressed and shown to act as a typical 1,5-dienoyl-CoA hydratase that does not accept 1-enoyl-CoA. A gene previously assigned to a 1-enoyl-CoA hydratase was heterologously expressed and identified to code for a bifunctional crotonase/3-OH-butyryl-CoA dehydrogenase. In summary, the results consistently provide biochemical evidence that F. placidus and probably other archaea predominantly degrade aromatics via the Thauera/Azoarcus type and not or only to a minor extent via the predicted R. palustris-type benzoyl-CoA degradation pathway.


Assuntos
Acil Coenzima A/metabolismo , Archaeoglobales/enzimologia , Redes e Vias Metabólicas/fisiologia , Anaerobiose , Archaeoglobales/genética , Coenzima A/metabolismo , Enoil-CoA Hidratase/metabolismo , Compostos Férricos/metabolismo , Hidroliases/metabolismo , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/metabolismo , Thauera/metabolismo
7.
Appl Environ Microbiol ; 81(3): 1003-12, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25416759

RESUMO

Geoglobus acetivorans is a hyperthermophilic anaerobic euryarchaeon of the order Archaeoglobales isolated from deep-sea hydrothermal vents. A unique physiological feature of the members of the genus Geoglobus is their obligate dependence on Fe(III) reduction, which plays an important role in the geochemistry of hydrothermal systems. The features of this organism and its complete 1,860,815-bp genome sequence are described in this report. Genome analysis revealed pathways enabling oxidation of molecular hydrogen, proteinaceous substrates, fatty acids, aromatic compounds, n-alkanes, and organic acids, including acetate, through anaerobic respiration linked to Fe(III) reduction. Consistent with the inability of G. acetivorans to grow on carbohydrates, the modified Embden-Meyerhof pathway encoded by the genome is incomplete. Autotrophic CO2 fixation is enabled by the Wood-Ljungdahl pathway. Reduction of insoluble poorly crystalline Fe(III) oxide depends on the transfer of electrons from the quinone pool to multiheme c-type cytochromes exposed on the cell surface. Direct contact of the cells and Fe(III) oxide particles could be facilitated by pilus-like appendages. Genome analysis indicated the presence of metabolic pathways for anaerobic degradation of aromatic compounds and n-alkanes, although an ability of G. acetivorans to grow on these substrates was not observed in laboratory experiments. Overall, our results suggest that Geoglobus species could play an important role in microbial communities of deep-sea hydrothermal vents as lithoautotrophic producers. An additional role as decomposers would close the biogeochemical cycle of carbon through complete mineralization of various organic compounds via Fe(III) respiration.


Assuntos
Acetatos/metabolismo , Archaeoglobales/genética , Processos Autotróficos , Compostos Ferrosos/metabolismo , Genoma Arqueal , Redes e Vias Metabólicas , Hidrocarbonetos Policíclicos Aromáticos/metabolismo , Biotransformação , DNA Arqueal/química , DNA Arqueal/genética , Dados de Sequência Molecular , Oxirredução , Análise de Sequência de DNA
8.
Microbiology (Reading) ; 160(Pt 12): 2694-2709, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25269449

RESUMO

Ferroglobus placidus was discovered to oxidize completely the aromatic amino acids tyrosine, phenylalanine and tryptophan when Fe(III) oxide was provided as an electron acceptor. This property had not been reported previously for a hyperthermophilic archaeon. It appeared that F. placidus follows a pathway for phenylalanine and tryptophan degradation similar to that of mesophilic nitrate-reducing bacteria, Thauera aromatica and Aromatoleum aromaticum EbN1. Phenylacetate, 4-hydroxyphenylacetate and indole-3-acetate were formed during anaerobic degradation of phenylalanine, tyrosine and tryptophan, respectively. Candidate genes for enzymes involved in the anaerobic oxidation of phenylalanine to phenylacetate (phenylalanine transaminase, phenylpyruvate decarboxylase and phenylacetaldehyde : ferredoxin oxidoreductase) were identified in the F. placidus genome. In addition, transcription of candidate genes for the anaerobic phenylacetate degradation, benzoyl-CoA degradation and glutaryl-CoA degradation pathways was significantly upregulated in microarray and quantitative real-time-PCR studies comparing phenylacetate-grown cells with acetate-grown cells. These results suggested that the general strategies for anaerobic degradation of aromatic amino acids are highly conserved amongst bacteria and archaea living in both mesophilic and hyperthermophilic environments. They also provided insights into the diverse metabolism of Archaeoglobaceae species living in hyperthermophilic environments.


Assuntos
Aminoácidos Aromáticos/metabolismo , Archaeoglobales/metabolismo , Anaerobiose , Biotransformação , Perfilação da Expressão Gênica , Ácidos Indolacéticos/metabolismo , Redes e Vias Metabólicas/genética , Análise em Microsséries , Dados de Sequência Molecular , Oxirredução , Fenilacetatos/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Análise de Sequência de DNA
9.
Appl Environ Microbiol ; 79(15): 4694-700, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23728807

RESUMO

The microbial reduction of Fe(III) plays an important role in the geochemistry of hydrothermal systems, yet it is poorly understood at the mechanistic level. Here we show that the obligate Fe(III)-reducing archaeon Geoglobus ahangari uses a direct-contact mechanism for the reduction of Fe(III) oxides to magnetite at 85°C. Alleviating the need to directly contact the mineral with the addition of a chelator or the electron shuttle anthraquinone-2,6-disulfonate (AQDS) stimulated Fe(III) reduction. In contrast, entrapment of the oxides within alginate beads to prevent cell contact with the electron acceptor prevented Fe(III) reduction and cell growth unless AQDS was provided. Furthermore, filtered culture supernatant fluids had no effect on Fe(III) reduction, ruling out the secretion of an endogenous mediator too large to permeate the alginate beads. Consistent with a direct contact mechanism, electron micrographs showed cells in intimate association with the Fe(III) mineral particles, which once dissolved revealed abundant curled appendages. The cells also produced several heme-containing proteins. Some of them were detected among proteins sheared from the cell's outer surface and were required for the reduction of insoluble Fe(III) oxides but not for the reduction of the soluble electron acceptor Fe(III) citrate. The results thus support a mechanism in which the cells directly attach and transfer electrons to the Fe(III) oxides using redox-active proteins exposed on the cell surface. This strategy confers on G. ahangari a competitive advantage for accessing and reducing Fe(III) oxides under the extreme physical and chemical conditions of hot ecosystems.


Assuntos
Antraquinonas/metabolismo , Archaeoglobales/metabolismo , Transporte de Elétrons , Compostos Férricos/metabolismo , Temperatura Alta , México , Oxirredução
10.
PLoS One ; 8(3): e59927, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23555835

RESUMO

To characterize high-temperature cellulolytic microbial communities, two lignocellulosic substrates, ammonia fiber-explosion-treated corn stover and aspen shavings, were incubated at average temperatures of 77 and 85°C in the sediment and water column of Great Boiling Spring, Nevada. Comparison of 109,941 quality-filtered 16S rRNA gene pyrosequences (pyrotags) from eight enrichments to 37,057 quality-filtered pyrotags from corresponding natural samples revealed distinct enriched communities dominated by phylotypes related to cellulolytic and hemicellulolytic Thermotoga and Dictyoglomus, cellulolytic and sugar-fermenting Desulfurococcales, and sugar-fermenting and hydrogenotrophic Archaeoglobales. Minor enriched populations included close relatives of hydrogenotrophic Thermodesulfobacteria, the candidate bacterial phylum OP9, and candidate archaeal groups C2 and DHVE3. Enrichment temperature was the major factor influencing community composition, with a negative correlation between temperature and richness, followed by lignocellulosic substrate composition. This study establishes the importance of these groups in the natural degradation of lignocellulose at high temperatures and suggests that a substantial portion of the diversity of thermophiles contributing to consortial cellulolysis may be contained within lineages that have representatives in pure culture.


Assuntos
Genes Arqueais , Fontes Termais/microbiologia , Lignina/química , Consórcios Microbianos/genética , Análise de Sequência de DNA/métodos , Archaeoglobales/genética , Biodiversidade , Desulfurococcales/genética , Temperatura Alta , Nevada , Filogenia , Polissacarídeos/química , RNA Ribossômico 16S/metabolismo , Árvores/microbiologia , Água/química , Microbiologia da Água , Zea mays/microbiologia
11.
Acta Crystallogr Sect F Struct Biol Cryst Commun ; 67(Pt 11): 1414-6, 2011 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-22102245

RESUMO

The cytidine at the first anticodon position of archaeal tRNA(Ile2), which decodes the isoleucine AUA codon, is modified to 2-agmatinylcytidine (agm(2)C) to guarantee the fidelity of protein biosynthesis. This post-transcriptional modification is catalyzed by tRNA(Ile)-agm(2)C synthetase (TiaS) using ATP and agmatine as substrates. Archaeoglobus fulgidus TiaS was overexpressed in Escherichia coli cells and purified. tRNA(Ile2) was prepared by in vitro transcription with T7 RNA polymerase. TiaS was cocrystallized with both tRNA(Ile2) and ATP by the vapour-diffusion method. The crystals of the TiaS-tRNA(Ile2)-ATP complex diffracted to 2.9 Å resolution using synchrotron radiation at the Photon Factory. The crystals belonged to the primitive hexagonal space group P3(2)21, with unit-cell parameters a = b = 131.1, c = 86.6 Å. The asymmetric unit is expected to contain one TiaS-tRNA(Ile2)-ATP complex, with a Matthews coefficient of 2.8 Å(3) Da(-1) and a solvent content of 61%.


Assuntos
Trifosfato de Adenosina/química , Archaeoglobales/enzimologia , Isoleucina-tRNA Ligase/química , RNA de Transferência de Isoleucina/química , Trifosfato de Adenosina/metabolismo , Cristalização , Cristalografia por Raios X , Isoleucina-tRNA Ligase/metabolismo , Ligação Proteica , RNA de Transferência de Isoleucina/metabolismo
12.
Appl Environ Microbiol ; 77(17): 5926-33, 2011 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-21742914

RESUMO

Anaerobic benzene oxidation coupled to the reduction of Fe(III) was studied in Ferroglobus placidus in order to learn more about how such a stable molecule could be metabolized under strict anaerobic conditions. F. placidus conserved energy to support growth at 85°C in a medium with benzene provided as the sole electron donor and Fe(III) as the sole electron acceptor. The stoichiometry of benzene loss and Fe(III) reduction, as well as the conversion of [(14)C]benzene to [(14)C]carbon dioxide, was consistent with complete oxidation of benzene to carbon dioxide with electron transfer to Fe(III). Benzoate, but not phenol or toluene, accumulated at low levels during benzene metabolism, and [(14)C]benzoate was produced from [(14)C]benzene. Analysis of gene transcript levels revealed increased expression of genes encoding enzymes for anaerobic benzoate degradation during growth on benzene versus growth on acetate, but genes involved in phenol degradation were not upregulated during growth on benzene. A gene for a putative carboxylase that was more highly expressed in benzene- than in benzoate-grown cells was identified. These results suggest that benzene is carboxylated to benzoate and that phenol is not an important intermediate in the benzene metabolism of F. placidus. This is the first demonstration of a microorganism in pure culture that can grow on benzene under strict anaerobic conditions and for which there is strong evidence for degradation of benzene via clearly defined anaerobic metabolic pathways. Thus, F. placidus provides a much-needed pure culture model for further studies on the anaerobic activation of benzene in microorganisms.


Assuntos
Archaeoglobales/metabolismo , Benzeno/metabolismo , Anaerobiose , Radioisótopos de Carbono/metabolismo , Compostos Férricos/metabolismo , Perfilação da Expressão Gênica , Temperatura Alta , Marcação por Isótopo , Oxirredução
13.
Protein Pept Lett ; 16(10): 1201-6, 2009.
Artigo em Inglês | MEDLINE | ID: mdl-19508182

RESUMO

In the last years we have performed a series of studies to characterize the conformational stability of three esterases from thermophilic and mesophilic sources: Aes esterase from Escherichia coli, EST2 from Alicyclobacillus acidocaldarius and AFEST from Archeoglobus fulgidus. These three esterases belong to the Hormone-sensitive lipase group of the superfamily of carboxylester hydrolases. The conformational stability of the three enzymes against temperature, urea and GuHCl has been determined by means of circular dichroism, fluorescence and differential scanning calorimetry measurements. Analysis of experimental data coupled with available structural information allowed us to suggest that the optimization of charge-charge interactions on the protein surface could one of the mechanisms to increase the thermal stability for the three esterases. This idea has been tested in the case of EST2, which shows a fully reversible thermal unfolding, by producing and studying variant forms of wild type enzyme in which a charged residue has been mutated. In the present article the obtained results are critically recollected in order to provide a clear and unified scenario.


Assuntos
Proteínas Arqueais/química , Proteínas de Bactérias/química , Esterases/química , Conformação Proteica , Alicyclobacillus/enzimologia , Alicyclobacillus/genética , Proteínas Arqueais/genética , Proteínas Arqueais/metabolismo , Archaeoglobales/enzimologia , Archaeoglobales/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Hidrolases de Éster Carboxílico/química , Hidrolases de Éster Carboxílico/genética , Hidrolases de Éster Carboxílico/metabolismo , Domínio Catalítico , Dicroísmo Circular , Cristalografia por Raios X , Estabilidade Enzimática , Escherichia coli/enzimologia , Escherichia coli/genética , Esterases/genética , Esterases/metabolismo , Guanidina/química , Cinética , Mutação , Dobramento de Proteína , Estrutura Terciária de Proteína , Esterol Esterase/química , Esterol Esterase/genética , Esterol Esterase/metabolismo , Temperatura , Ureia/química
14.
J Bacteriol ; 191(1): 375-87, 2009 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-18952801

RESUMO

HAMP domains, found in many bacterial signal transduction proteins, generally transmit an intramolecular signal between an extracellular sensory domain and an intracellular signaling domain. Studies of HAMP domains in proteins where both the input and output signals occur intracellularly are limited to those of the Aer energy taxis receptor of Escherichia coli, which has both a HAMP domain and a sensory PAS domain. Campylobacter jejuni has an energy taxis system consisting of the domains of Aer divided between two proteins, CetA (HAMP domain containing) and CetB (PAS domain containing). In this study, we found that the CetA HAMP domain differs significantly from that of Aer in the predicted secondary structure. Using similarity searches, we identified 55 pairs of HAMP/PAS proteins encoded by adjacent genes in a diverse group of microorganisms. We propose that these HAMP/PAS pairs form a new family of bipartite energy taxis receptors. Within these proteins, we identified nine residues in the HAMP domain and proximal signaling domain that are highly conserved, at least three of which are required for CetA function. Additionally, we demonstrated that CetA contributes to the invasion of human epithelial cells by C. jejuni, while CetB does not. This finding supports the hypothesis that members of HAMP/PAS pairs possess the capacity to act independently of each other in cellular traits other than energy taxis.


Assuntos
Campylobacter jejuni/genética , Proteínas de Transporte/genética , Sequência de Aminoácidos , Proteínas Arqueais/química , Proteínas Arqueais/genética , Proteínas Arqueais/metabolismo , Archaeoglobales/genética , Campylobacter jejuni/metabolismo , Campylobacter jejuni/patogenicidade , Proteínas de Transporte/química , Proteínas de Transporte/metabolismo , Linhagem Celular , Quimiotaxia/genética , Sequência Conservada , Células Epiteliais/microbiologia , Células Epiteliais/fisiologia , Escherichia coli/genética , Escherichia coli/metabolismo , Escherichia coli/patogenicidade , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Humanos , Dados de Sequência Molecular , Estrutura Secundária de Proteína , Transdução de Sinais/genética
15.
Adv Space Res ; 35(9): 1634-42, 2005.
Artigo em Inglês | MEDLINE | ID: mdl-16175703

RESUMO

A sub-surface archaeal community at the Suiyo Seamount in the Western Pacific Ocean was investigated by 16S rRNA gene sequence and whole-cell in situ hybridization analyses. In this study, we drilled and cased holes at the hydrothermal area of the seamount to minimize contamination of the hydrothermal fluid in the sub-seafloor by penetrating seawater. PCR clone analysis of the hydrothermal fluid samples collected from a cased hole indicated the presence of chemolithoautotrophic primary biomass producers of Archaeoglobales and the Methanococcales-related archaeal HTE1 group, both of which can utilize hydrogen as an electron donor. We discuss the implication of the microbial community on the early history of life and on the search for extraterrestrial life.


Assuntos
Archaeoglobales/isolamento & purificação , Ecossistema , Methanococcales/isolamento & purificação , RNA Ribossômico 16S/genética , Água do Mar/microbiologia , Archaea/genética , Archaeoglobales/genética , Biomassa , Temperatura Alta , Japão , Methanococcales/genética , Oceano Pacífico , Filogenia , Reação em Cadeia da Polimerase , RNA Arqueal , Microbiologia da Água
16.
J Bacteriol ; 187(17): 6046-57, 2005 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-16109946

RESUMO

The heat shock response of the hyperthermophilic archaeon Archaeoglobus fulgidus strain VC-16 was studied using whole-genome microarrays. On the basis of the resulting expression profiles, approximately 350 of the 2,410 open reading frames (ORFs) (ca. 14%) exhibited increased or decreased transcript abundance. These span a range of cell functions, including energy production, amino acid metabolism, and signal transduction, where the majority are uncharacterized. One ORF called AF1298 was identified that contains a putative helix-turn-helix DNA binding motif. The gene product, HSR1, was expressed and purified from Escherichia coli and was used to characterize specific DNA recognition regions upstream of two A. fulgidus genes, AF1298 and AF1971. The results indicate that AF1298 is autoregulated and is part of an operon with two downstream genes that encode a small heat shock protein, Hsp20, and cdc48, an AAA+ ATPase. The DNase I footprints using HSR1 suggest the presence of a cis-binding motif upstream of AF1298 consisting of CTAAC-N5-GTTAG. Since AF1298 is negatively regulated in response to heat shock and encodes a protein only distantly related to the N-terminal DNA binding domain of Phr of Pyrococcus furiosus, these results suggest that HSR1 and Phr may belong to an evolutionarily diverse protein family involved in heat shock regulation in hyperthermophilic and mesophilic Archaea organisms.


Assuntos
Proteínas Arqueais/genética , Archaeoglobales/genética , Proteínas de Choque Térmico/genética , Sequência de Aminoácidos , Sequência Conservada , Regulação da Expressão Gênica em Archaea , Cinética , Dados de Sequência Molecular , Análise de Sequência com Séries de Oligonucleotídeos , Fases de Leitura Aberta , Reação em Cadeia da Polimerase , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Homologia de Sequência do Ácido Nucleico
17.
J Biol Chem ; 279(40): 41822-9, 2004 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-15286083

RESUMO

In Archaea, fibrillarin and Nop5p form the core complex of box C/D small ribonucleoprotein particles, which are responsible for site-specific 2'-hydroxyl methylation of ribosomal and transfer RNAs. Fibrillarin has a conserved methyltransferase fold and employs S-adenosyl-l-methionine (AdoMet) as the cofactor in methyl transfer reactions. Comparison between recently determined crystal structures of free fibrillarin and fibrillarin-Nop5p-AdoMet tertiary complex revealed large conformational differences at the cofactor-binding site in fibrillarin. To identify the structural elements responsible for these large conformational differences, we refined a crystal structure of Archaeoglobus fulgidus fibrillarin-Nop5p binary complex at 3.5 A. This structure exhibited a pre-formed backbone geometry at the cofactor binding site similar to that when the cofactor is bound, suggesting that binding of Nop5p alone to fibrillarin is sufficient to stabilize the AdoMet-binding pocket. Calorimetry studies of cofactor binding to fibrillarin alone and to fibrillarin-Nop5p binary complex provided further support for this role of Nop5p. Mutagenesis and thermodynamic data showed that a cation-pi bridge formed between Tyr-89 of fibrillarin and Arg-169 of Nop5p, although dispensable for in vitro methylation activity, could partially account for the enhanced binding of cofactor to fibrillarin by Nop5p. Finally, assessment of cofactor-binding thermodynamics and catalytic activities of enzyme mutants identified three additional fibrillarin residues (Thr-70, Glu-88, and Asp-133) to be important for cofactor binding and for catalysis.


Assuntos
Proteínas Arqueais/química , Proteínas Cromossômicas não Histona/metabolismo , Proteínas Nucleares/química , S-Adenosilmetionina/metabolismo , Termodinâmica , Archaeoglobales/química , Sítios de Ligação , Calorimetria , Proteínas Cromossômicas não Histona/química , Cristalografia por Raios X , Ligação de Hidrogênio , Metilação , Ligação Proteica , Ribonucleoproteínas , S-Adenosilmetionina/química
18.
FEBS Lett ; 540(1-3): 171-5, 2003 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-12681503

RESUMO

The AlkA protein from the archaebacterium Archaeglobus fulgidus was characterised with respect to release of hypoxanthine from DNA. The hypoxanthine glycosylase activity had optimal activity at 60 degrees C at pH 5.0. The enzyme released hypoxanthine from substrates with a preference for dI:dG >> dI:dT > dI:dC > dI:dA. The presence of a mismatch on either side of the dIMP in the substrate reduced excision efficiency of the hypoxanthine residue at neutral pH, while a mismatch on both sides of the dIMP resulted in total loss of excision. Release of hypoxanthine from DNA required a minimum of two bases on the 5' side and four bases on the 3' side of the dIMP residue.


Assuntos
Archaeoglobales/enzimologia , Glicosídeo Hidrolases/metabolismo , Pareamento Incorreto de Bases , Sequência de Bases , DNA Arqueal , Concentração de Íons de Hidrogênio , Hipoxantina/metabolismo
19.
Mol Biol Evol ; 18(7): 1378-88, 2001 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-11420376

RESUMO

The enzyme 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG-CoA reductase or HMGR) fulfills an essential role in archaea, as it is required for the synthesis of isoprenoid ethers, the main component of archaeal cell membranes. There are two clearly homologous but structurally different classes of the enzyme, one found mainly in eukaryotes and archaea (class 1), and the other found in bacteria (class 2). This feature facilitated the identification of several cases of interdomain lateral gene transfer (LGT), in particular, the bacterial origin for the HMGR gene from the archaeon Archaeoglobus fulgidus. In order to investigate if this LGT event was recent and limited in its scope or had a broad and long-term impact on the recipient and its related lineages, the HMGR gene was amplified and sequenced from a variety of archaea. The survey covered close relatives of A. fulgidus, the only archaeon known prior to this study to possess a bacterial-like HMGR; representatives of each main euryarchaeal group were also inspected. All culturable members of the archaeal group Archaeoglobales were found to display an HMGR very similar to the enzyme of the bacterium Pseudomonas mevalonii. Surprisingly, two species of the genus Thermoplasma also harbor an HMGR of bacterial origin highly similar to the enzymes found in the Archaeoglobales. Phylogenetic analyses of the HMGR gene and comparisons to reference phylogenies from other genes confirm a common bacterial origin for the HMGRs of Thermoplasmatales and Archaeoglobales. The most likely explanation of these results includes an initial bacteria-to-archaea transfer, followed by a another event between archaea. Their presence in two divergent archaeal lineages suggests an important adaptive role for these laterally transferred genes.


Assuntos
Archaeoglobales/enzimologia , Archaeoglobales/genética , Bactérias/enzimologia , Bactérias/genética , Hidroximetilglutaril-CoA Redutases/genética , Thermoplasmales/enzimologia , Thermoplasmales/genética , Archaeoglobales/classificação , Sequência de Bases , Primers do DNA/genética , Evolução Molecular , Transferência Genética Horizontal , Genes Arqueais , Dados de Sequência Molecular , Filogenia , Especificidade da Espécie , Thermoplasmales/classificação
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...