Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 1.752
Filtrar
1.
Extremophiles ; 25(5-6): 513-526, 2021 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-34647163

RESUMEN

Thermoanaerobacter kivui is an acetogenic model organism that reduces CO2 with electrons derived from H2 or CO, or from organic substrates in the Wood-Ljugdahl pathway (WLP). For the calculation of ATP yields, it is necessary to know the electron carriers involved in coupling of the oxidative and reductive parts of metabolism. Analyses of key catabolic oxidoreductases in cell-free extract (CFE) or with purified enzymes revealed the physiological electron carriers involved. The glyceraldehyde-3-phosphate dehydrogenase (GA3P-DH) assayed in CFE was NAD+-specific, NADP+ was used with less than 4% and ferredoxin (Fd) was not used. The methylene-THF dehydrogenase was NADP+-specific, NAD+ or Fd were not used. A Nfn-type transhydrogenase that catalyzes reduced Fd-dependent reduction of NADP+ with NADH as electron donor was also identified in CFE. The electron carriers used by the potential electron-bifurcating hydrogenase (HydABC) could not be unambiguously determined in CFE for technical reasons. Therefore, the enzyme was produced homologously in T. kivui and purified by affinity chromatography. HydABC contained 33.9 ± 4.5 mol Fe/mol of protein and FMN; it reduced NADP+ but not NAD+. The methylene-THF reductase (MetFV) was also produced homologously in T. kivui and purified by affinity chromatography. MetFV contained 7.2 ± 0.4 mol Fe/mol of protein and FMN; the complex did neither use NADPH nor NADH as reductant but only reduced Fd. In sum, these analysis allowed us to propose a scheme for entire electron flow and bioenergetics in T. kivui.


Asunto(s)
Electrones , Hidrogenasas , Procesos Autotróficos , Hidrogenasas/metabolismo , NAD/metabolismo , NADP , Oxidación-Reducción , Thermoanaerobacter/metabolismo
2.
Nat Commun ; 12(1): 5281, 2021 09 06.
Artículo en Inglés | MEDLINE | ID: mdl-34489402

RESUMEN

The archaeal phylum Woesearchaeota, within the DPANN superphylum, includes phylogenetically diverse microorganisms that inhabit various environments. Their biology is poorly understood due to the lack of cultured isolates. Here, we analyze datasets of Woesearchaeota 16S rRNA gene sequences and metagenome-assembled genomes to infer global distribution patterns, ecological preferences and metabolic capabilities. Phylogenomic analyses indicate that the phylum can be classified into ten subgroups, termed A-J. While a symbiotic lifestyle is predicted for most, some members of subgroup J might be host-independent. The genomes of several Woesearchaeota, including subgroup J, encode putative [FeFe] hydrogenases (known to be important for fermentation in other organisms), suggesting that these archaea might be anaerobic fermentative heterotrophs.


Asunto(s)
Archaea/genética , Proteínas Arqueales/genética , Genoma Arqueal , Hidrogenasas/genética , ARN de Archaea/genética , ARN Ribosómico 16S/genética , Secuencia de Aminoácidos , Anaerobiosis/genética , Archaea/clasificación , Archaea/enzimología , Proteínas Arqueales/metabolismo , Evolución Biológica , Fermentación , Procesos Heterotróficos/genética , Hidrogenasas/metabolismo , Metagenoma , Filogenia , Alineación de Secuencia , Homología de Secuencia de Aminoácido
3.
Angew Chem Int Ed Engl ; 60(38): 20744-20747, 2021 09 13.
Artículo en Inglés | MEDLINE | ID: mdl-34324230

RESUMEN

Azadithiolate, a cofactor found in all [FeFe]-hydrogenases, is shown to undergo acid-catalyzed rearrangement. Fe2 [(SCH2 )2 NH](CO)6 self-condenses to give Fe6 [(SCH2 )3 N]2 (CO)17 . The reaction, which is driven by loss of NH4 + , illustrates the exchange of the amine group. X-ray crystallography reveals that three Fe2 (SR)2 (CO)x butterfly subunits interconnected by the aminotrithiolate [N(CH2 S)3 ]3- . Mechanistic studies reveal that Fe2 [(SCH2 )2 NR](CO)6 participate in a range of amine exchange reactions, enabling new methodologies for modifying the adt cofactor. Ru2 [(SCH2 )2 NH](CO)6 also rearranges, but proceeds further to give derivatives with Ru-alkyl bonds Ru6 [(SCH2 )3 N][(SCH2 )2 NCH2 ]S(CO)17 and [Ru2 [(SCH2 )2 NCH2 ](CO)5 ]2 S.


Asunto(s)
Compuestos Aza/metabolismo , Complejos de Coordinación/metabolismo , Hidrogenasas/metabolismo , Rubidio/metabolismo , Tolueno/análogos & derivados , Compuestos Aza/química , Complejos de Coordinación/química , Modelos Moleculares , Estructura Molecular , Rubidio/química , Tolueno/química , Tolueno/metabolismo
4.
Elife ; 102021 06 04.
Artículo en Inglés | MEDLINE | ID: mdl-34085924

RESUMEN

The composition of gut-associated microbial communities changes during intestinal inflammation, including an expansion of Enterobacteriaceae populations. The mechanisms underlying microbiota changes during inflammation are incompletely understood. Here, we analyzed previously published metagenomic datasets with a focus on microbial hydrogen metabolism. The bacterial genomes in the inflamed murine gut and in patients with inflammatory bowel disease contained more genes encoding predicted hydrogen-utilizing hydrogenases compared to communities found under non-inflamed conditions. To validate these findings, we investigated hydrogen metabolism of Escherichia coli, a representative Enterobacteriaceae, in mouse models of colitis. E. coli mutants lacking hydrogenase-1 and hydrogenase-2 displayed decreased fitness during colonization of the inflamed cecum and colon. Utilization of molecular hydrogen was in part dependent on respiration of inflammation-derived electron acceptors. This work highlights the contribution of hydrogenases to alterations of the gut microbiota in the context of non-infectious colitis.


Asunto(s)
Ciego/microbiología , Colitis/inducido químicamente , Colitis/microbiología , Colon/microbiología , Infecciones por Escherichia coli/microbiología , Escherichia coli/metabolismo , Microbioma Gastrointestinal , Hidrógeno/metabolismo , Animales , Ciego/metabolismo , Ciego/patología , Colitis/metabolismo , Colitis/patología , Colon/metabolismo , Colon/patología , Bases de Datos Genéticas , Sulfato de Dextran , Modelos Animales de Enfermedad , Disbiosis , Escherichia coli/genética , Escherichia coli/crecimiento & desarrollo , Infecciones por Escherichia coli/metabolismo , Infecciones por Escherichia coli/patología , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Femenino , Humanos , Hidrogenasas/genética , Hidrogenasas/metabolismo , Interleucina-10/genética , Interleucina-10/metabolismo , Masculino , Metagenoma , Metagenómica , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Piroxicam
5.
Parasitol Res ; 120(6): 2189-2198, 2021 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-33855618

RESUMEN

Recent phylogenetic and morphologic studies of Trichomonas spp. suggests that there are more than 3 species that infect the upper alimentary tract of wild birds, which include T. gallinae, T. stableri, and T. gypaetinii. In this study, investigations were conducted on the prevalence of trichomonads in the upper alimentary tract of 12 Steller's sea eagles (Haliaeetus pelagicus) and 18 white-tailed sea eagles (H. albicilla). All birds were rescued from the wild and kept at a rehabilitation facility in Hokkaido, Japan, for variable durations and did not show any symptoms of trichomonosis. The ITS1-5.8SrRNA-ITS2 (ITS) genomic region of Trichomonas spp. was detected from 29 samples by PCR, and flagellates were confirmed from 4 samples by culture. Morphologic observations and measurement recordings were conducted under a light microscope on trophozoites obtained from the cultured isolates. Genomic sequences of the ITS, 18S ribosomal RNA (18S rRNA), Fe-hydrogenase, and RNA polymerase II largest subunit (Rpb1) regions were determined by direct sequencing, and phylogenetic analyses were conducted with previously published sequences of Trichomonas spp. All isolates were concluded as T. gypaetinii based on morphologic and molecular characterizations of the ITS and 18S rRNA genes. This is the first study to isolate T. gypaetinii from Haliaeetus eagles and further provide novel sequences of the Fe-hydrogenase and Rpb1 genes of T. gypaetinii. Both genomic regions also confirmed that T. gypaetinii belong to independent clusters from other Trichomonas spp.


Asunto(s)
Enfermedades de las Aves/parasitología , Águilas/parasitología , Tricomoniasis/veterinaria , Animales , Animales Salvajes/parasitología , Enfermedades de las Aves/epidemiología , Águilas/genética , Femenino , Hidrogenasas/metabolismo , Proteínas Hierro-Azufre/metabolismo , Japón/epidemiología , Masculino , Reacción en Cadena de la Polimerasa/veterinaria , Trichomonas , Tricomoniasis/epidemiología , Tricomoniasis/parasitología
6.
J Biol Chem ; 296: 100710, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33930466

RESUMEN

Pyrococcus furiosus is a hyperthermophilic anaerobic archaeon whose metabolism depends on whether elemental sulfur is (+S0) or is not (-S0) included in growth medium. Under +S0 conditions, expression of respiratory hydrogenase declines while respiratory membrane-bound sulfane reductase and the putative iron-storage protein IssA increase. Our objective was to investigate the iron content of WT and ΔIssA cells under these growth conditions using Mössbauer spectroscopy. WT-S0 cells contained ∼1 mM Fe, with ∼85% present as two spectroscopically distinct forms of S = 0 [Fe4S4]2+ clusters; the remainder was mainly high-spin FeII. WT+S0 cells contained 5 to 9 mM Fe, with 75 to 90% present as magnetically ordered thioferrate-like (TFL) iron nanoparticles. TFL iron was similar to chemically defined thioferrates; both consisted of FeIII ions coordinated by an S4 environment, and both exhibited strong coupling between particles causing high applied fields to have little spectral effect. At high temperatures with magnetic hyperfine interactions abolished, TFL iron exhibited two doublets overlapping those of [Fe4S4]2+ clusters in -S0 cells. This coincidence arose because of similar coordination environments of TFL iron and cluster iron. The TFL structure was more heterogeneous in the presence of IssA. Presented data suggest that IssA may coordinate insoluble iron sulfides as TFL iron, formed as a byproduct of anaerobic sulfur respiration under high iron conditions, which thereby reduces its toxicity to the cell. This was the first Mössbauer characterization of the ironome of an archaeon, and it illustrates differences relative to the iron content of better-studied bacteria such as Escherichia coli.


Asunto(s)
Hierro/metabolismo , Pyrococcus furiosus/metabolismo , Azufre/metabolismo , Hidrogenasas/metabolismo , Oxidación-Reducción
7.
Physiol Plant ; 173(2): 555-567, 2021 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-33860946

RESUMEN

Photosynthetic production of molecular hydrogen (H2 ) by cyanobacteria and green algae is a potential source of renewable energy. These organisms are capable of water biophotolysis by taking advantage of photosynthetic apparatus that links water oxidation at Photosystem II and reduction of protons to H2 downstream of Photosystem I. Although the process has a theoretical potential to displace fossil fuels, photosynthetic H2 production in its current state is not yet efficient enough for industrial applications due to a number of physiological, biochemical, and engineering barriers. This article presents a short overview of the metabolic pathways and enzymes involved in H2 photoproduction in cyanobacteria and green algae and our present understanding of the mechanisms of this process. We also summarize recent advances in engineering photosynthetic cell factories capable of overcoming the major barriers to efficient and sustainable H2 production.


Asunto(s)
Chlorophyta , Hidrogenasas , Chlorophyta/genética , Chlorophyta/metabolismo , Hidrógeno , Hidrogenasas/genética , Hidrogenasas/metabolismo , Fotosíntesis , Complejo de Proteína del Fotosistema II/metabolismo
8.
Angew Chem Int Ed Engl ; 60(26): 14488-14497, 2021 06 21.
Artículo en Inglés | MEDLINE | ID: mdl-33871139

RESUMEN

An artificial amyloid-based redox hydrogel was designed for mediating electron transfer between a [NiFeSe] hydrogenase and an electrode. Starting from a mutated prion-forming domain of fungal protein HET-s, a hybrid redox protein containing a single benzyl methyl viologen moiety was synthesized. This protein was able to self-assemble into structurally homogenous nanofibrils. Molecular modeling confirmed that the redox groups are aligned along the fibril axis and are tethered to its core by a long, flexible polypeptide chain that allows close encounters between the fibril-bound oxidized or reduced redox groups. Redox hydrogel films capable of immobilizing the hydrogenase under mild conditions at the surface of carbon electrodes were obtained by a simple pH jump. In this way, bioelectrodes for the electrocatalytic oxidation of H2 were fabricated that afforded catalytic current densities of up to 270 µA cm-2 , with an overpotential of 0.33 V, under quiescent conditions at 45 °C.


Asunto(s)
Amiloide/metabolismo , Hidrogeles/metabolismo , Hidrógeno/metabolismo , Hidrogenasas/metabolismo , Amiloide/química , Biocatálisis , Electrodos , Transporte de Electrón , Hidrogeles/química , Hidrógeno/química , Hidrogenasas/química , Modelos Moleculares , Oxidación-Reducción , Tamaño de la Partícula
9.
Angew Chem Int Ed Engl ; 60(25): 13824-13828, 2021 06 14.
Artículo en Inglés | MEDLINE | ID: mdl-33721401

RESUMEN

A new activity for the [NiFe] uptake hydrogenase 1 of Escherichia coli (Hyd1) is presented. Direct reduction of biological flavin cofactors FMN and FAD is achieved using H2 as a simple, completely atom-economical reductant. The robust nature of Hyd1 is exploited for flavin reduction across a broad range of temperatures (25-70 °C) and extended reaction times. The utility of this system as a simple, easy to implement FMNH2 or FADH2 regenerating system is then demonstrated by supplying reduced flavin to Old Yellow Enzyme "ene-reductases" to support asymmetric alkene reductions with up to 100 % conversion. Hyd1 turnover frequencies up to 20.4 min-1 and total turnover numbers up to 20 200 were recorded during flavin recycling.


Asunto(s)
Alquenos/metabolismo , Escherichia coli/enzimología , Flavinas/metabolismo , Hidrogenasas/metabolismo , Oxidorreductasas/metabolismo , Alquenos/química , Biocatálisis , Flavinas/química , Hidrogenasas/química , Hidrogenación , Estructura Molecular , Oxidación-Reducción , Oxidorreductasas/química
10.
Biochim Biophys Acta Bioenerg ; 1862(6): 148401, 2021 06 01.
Artículo en Inglés | MEDLINE | ID: mdl-33684340

RESUMEN

The concomitant presence of two distinctive polypeptide modules, which we have chosen to denominate as the "Y-junction" and the "flavin" module, is observed in 3D structures of enzymes as functionally diverse as complex I, NAD(P)-dependent [NiFe]-hydrogenases and NAD(P)-dependent formate dehydrogenases. Amino acid sequence conservation furthermore suggests that both modules are also part of NAD(P)-dependent [FeFe]-hydrogenases for which no 3D structure model is available yet. The flavin module harbours the site of interaction with the substrate NAD(P) which exchanges two electrons with a strictly conserved flavin moiety. The Y-junction module typically contains four iron-sulphur centres arranged to form a Y-shaped electron transfer conduit and mediates electron transfer between the flavin module and the catalytic units of the respective enzymes. The Y-junction module represents an electron transfer hub with three potential electron entry/exit sites. The pattern of specific redox centres present both in the Y-junction and the flavin module is correlated to present knowledge of these enzymes' functional properties. We have searched publicly accessible genomes for gene clusters containing both the Y-junction and the flavin module to assemble a comprehensive picture of the diversity of enzymes harbouring this dyad of modules and to reconstruct their phylogenetic relationships. These analyses indicate the presence of the dyad already in the last universal common ancestor and the emergence of complex I's EFG-module out of a subgroup of NAD(P)- dependent formate dehydrogenases.


Asunto(s)
Proteínas Bacterianas/metabolismo , Electrones , Flavinas/metabolismo , Formiato Deshidrogenasas/metabolismo , Hidrogenasas/metabolismo , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Transporte de Electrón , Flavinas/química , Formiato Deshidrogenasas/química , Formiato Deshidrogenasas/genética , Hidrogenasas/química , Hidrogenasas/genética , Filogenia
11.
Angew Chem Int Ed Engl ; 60(29): 15854-15862, 2021 07 12.
Artículo en Inglés | MEDLINE | ID: mdl-33783938

RESUMEN

To study metalloenzymes in detail, we developed a new experimental setup allowing the controlled preparation of catalytic intermediates for characterization by various spectroscopic techniques. The in situ monitoring of redox transitions by infrared spectroscopy in enzyme lyophilizate, crystals, and solution during gas exchange in a wide temperature range can be accomplished as well. Two O2 -tolerant [NiFe]-hydrogenases were investigated as model systems. First, we utilized our platform to prepare highly concentrated hydrogenase lyophilizate in a paramagnetic state harboring a bridging hydride. This procedure proved beneficial for 57 Fe nuclear resonance vibrational spectroscopy and revealed, in combination with density functional theory calculations, the vibrational fingerprint of this catalytic intermediate. The same in situ IR setup, combined with resonance Raman spectroscopy, provided detailed insights into the redox chemistry of enzyme crystals, underlining the general necessity to complement X-ray crystallographic data with spectroscopic analyses.


Asunto(s)
Hidrogenasas/química , Hidrogenasas/metabolismo , Solventes/química , Dominio Catalítico , Cristalografía por Rayos X , Liofilización , Modelos Moleculares , Oxidación-Reducción
12.
ChemSusChem ; 14(10): 2237-2246, 2021 May 20.
Artículo en Inglés | MEDLINE | ID: mdl-33787007

RESUMEN

Hydrogenase enzymes produce H2 gas, which can be a potential source of alternative energy. Inspired by the [NiFe] hydrogenases, we report the construction of a de novo-designed artificial hydrogenase (ArH). The ArH is a dimeric coiled coil where two cysteine (Cys) residues are introduced at tandem a/d positions of a heptad to create a tetrathiolato Ni binding site. Spectroscopic studies show that Ni binding significantly stabilizes the peptide producing electronic transitions characteristic of Ni-thiolate proteins. The ArH produces H2 photocatalytically, demonstrating a bell-shaped pH-dependence on activity. Fluorescence lifetimes and transient absorption spectroscopic studies are undertaken to elucidate the nature of pH-dependence, and to monitor the reaction kinetics of the photochemical processes. pH titrations are employed to determine the role of protonated Cys on reactivity. Through combining these results, a fine balance is found between solution acidity and the electron transfer steps. This balance is critical to maximize the production of NiI -peptide and protonation of the NiII -H- intermediate (Ni-R) by a Cys (pKa ≈6.4) to produce H2 .


Asunto(s)
Materiales Biomiméticos/química , Cisteína/química , Diseño de Fármacos , Hidrogenasas/metabolismo , Procesos Fotoquímicos , Protones
13.
J Phys Chem A ; 125(7): 1413-1423, 2021 Feb 25.
Artículo en Inglés | MEDLINE | ID: mdl-33567824

RESUMEN

Two anionic asymmetric Fe-Fe hydrogenase model compounds containing a single cyano (CN) and five carboxyl (CO) ligands, [Et4N][Fe2(µ-S2C3H6)(CO)5(CN)1] and [Et4N][Fe2(µ-S2C2H4)(CO)5(CN)1], dissolved in room-temperature acetonitrile, are examined. The molecular asymmetry affects the redox potentials of the central iron atoms, thus changing the photophysics and possible catalytic properties of the compounds. Femtosecond ultraviolet excitation with mid-infrared probe spectroscopy of the model compounds was employed to better understand the ultrafast dynamics of the enzyme-active site. Continuous ultraviolet lamp excitation with Fourier transform infrared (FTIR) spectroscopy was also used to explore stable product formation on the second timescale. For both model compounds, two timescales are observed; a 20-30 ps decay and the formation of a long-lived photoproduct. The picosecond decay is assigned to vibrational cooling and rotational dynamics, while the residual spectra remain for up to 300 ps, suggesting the formation of new photoproducts. Static FTIR spectroscopy yielded a different stable photoproduct than that observed on the ultrafast timescale. Density functional theory calculations simulated photoproducts for CO-loss and CN-loss isomers, and the resulting photoproduct spectra suggest that the picosecond transients arise from a complex mixture of isomerization after CO-loss, while dimerization and formation of a CN-containing Fe-CO-Fe bridged species are also considered.


Asunto(s)
Proteínas Bacterianas/química , Cianuros/química , Hidrogenasas/química , Proteínas Hierro-Azufre/química , Proteínas Bacterianas/metabolismo , Catálisis , Dominio Catalítico , Cianuros/metabolismo , Compuestos Ferrosos/química , Compuestos Ferrosos/metabolismo , Hidrogenasas/metabolismo , Proteínas Hierro-Azufre/metabolismo , Modelos Químicos , Conformación Proteica , Espectrofotometría Infrarroja/métodos
14.
Angew Chem Int Ed Engl ; 60(24): 13350-13357, 2021 06 07.
Artículo en Inglés | MEDLINE | ID: mdl-33635597

RESUMEN

The reconstitution of [Mn]-hydrogenases using a series of MnI complexes is described. These complexes are designed to have an internal base or pro-base that may participate in metal-ligand cooperative catalysis or have no internal base or pro-base. Only MnI complexes with an internal base or pro-base are active for H2 activation; only [Mn]-hydrogenases incorporating such complexes are active for hydrogenase reactions. These results confirm the essential role of metal-ligand cooperation for H2 activation by the MnI complexes alone and by [Mn]-hydrogenases. Owing to the nature and position of the internal base or pro-base, the mode of metal-ligand cooperation in two active [Mn]-hydrogenases is different from that of the native [Fe]-hydrogenase. One [Mn]-hydrogenase has the highest specific activity of semi-synthetic [Mn]- and [Fe]-hydrogenases. This work demonstrates reconstitution of active artificial hydrogenases using synthetic complexes differing greatly from the native active site.


Asunto(s)
Complejos de Coordinación/química , Hidrogenasas/química , Ligandos , Manganeso/química , Materiales Biomiméticos/química , Materiales Biomiméticos/metabolismo , Catálisis , Dominio Catalítico , Hidrógeno/química , Hidrogenasas/metabolismo , Conformación Molecular
15.
Angew Chem Int Ed Engl ; 60(18): 10001-10006, 2021 04 26.
Artículo en Inglés | MEDLINE | ID: mdl-33630389

RESUMEN

Gas-processing metalloenzymes are of interest to future bio- and bioinspired technologies. Of particular importance are hydrogenases and nitrogenases, which both produce molecular hydrogen (H2 ) from proton (H+ ) reduction. Herein, we report on the use of rotating ring-disk electrochemistry (RRDE) and mass spectrometry (MS) to follow the production of H2 and isotopes produced from deuteron (D+ ) reduction (HD and D2 ) using the [FeFe]-hydrogenase from Clostridium pasteurianum, a model hydrogen-evolving metalloenzyme. This facilitates enzymology studies independent of non-innocent chemical reductants. We anticipate that these approaches will be of value in resolving the catalytic mechanisms of H2 -producing metalloenzymes and the design of bioinspired catalysts for H2 production and N2 fixation.


Asunto(s)
Hidrógeno/metabolismo , Hidrogenasas/metabolismo , Proteínas Hierro-Azufre/metabolismo , Clostridium/enzimología , Técnicas Electroquímicas , Electrodos , Hidrógeno/química , Hidrogenasas/química , Proteínas Hierro-Azufre/química , Espectrometría de Masas
16.
Dalton Trans ; 50(10): 3641-3650, 2021 Mar 16.
Artículo en Inglés | MEDLINE | ID: mdl-33629081

RESUMEN

Hydrogenases are bidirectional redox enzymes that catalyze hydrogen turnover in archaea, bacteria, and algae. While all types of hydrogenase show H2 oxidation activity, [FeFe]-hydrogenases are excellent H2 evolution catalysts as well. Their active site cofactor comprises a [4Fe-4S] cluster covalently linked to a diiron site equipped with carbon monoxide and cyanide ligands. The active site niche is connected with the solvent by two distinct proton transfer pathways. To analyze the catalytic mechanism of [FeFe]-hydrogenase, we employ operando infrared spectroscopy and infrared spectro-electrochemistry. Titrating the pH under H2 oxidation or H2 evolution conditions reveals the influence of site-selective protonation on the equilibrium of reduced cofactor states. Governed by pKa differences across the active site niche and proton transfer pathways, we find that individual electrons are stabilized either at the [4Fe-4S] cluster (alkaline pH values) or at the diiron site (acidic pH values). This observation is discussed in the context of the complex interdependence of hydrogen turnover and bulk pH.


Asunto(s)
Hidrogenasas/metabolismo , Proteínas Hierro-Azufre/metabolismo , Chlamydomonas reinhardtii/enzimología , Electrones , Concentración de Iones de Hidrógeno , Hidrogenasas/análisis , Proteínas Hierro-Azufre/análisis , Oxidación-Reducción , Protones
17.
Chem Commun (Camb) ; 57(14): 1750-1753, 2021 Feb 19.
Artículo en Inglés | MEDLINE | ID: mdl-33469641

RESUMEN

We demonstrate that the insertion of the dinuclear active site of [FeFe] hydrogenase into the apo-enzyme can occur when the enzyme is embedded in a film of redox polymer, under conditions of mediated electron transfer. The maturation can be monitored by electrochemistry, and is as fast as under conditions of direct electron transfer. This new approach further clears the way to the implementation of hydrogenases in large scale real life processes.


Asunto(s)
Enzimas Inmovilizadas/química , Enzimas Inmovilizadas/metabolismo , Hidrogenasas/química , Hidrogenasas/metabolismo , Proteínas Hierro-Azufre/química , Proteínas Hierro-Azufre/metabolismo , Membranas Artificiales , Polímeros/química
18.
ISME J ; 15(2): 363-376, 2021 02.
Artículo en Inglés | MEDLINE | ID: mdl-33024291

RESUMEN

Significant rates of atmospheric dihydrogen (H2) consumption have been observed in temperate soils due to the activity of high-affinity enzymes, such as the group 1h [NiFe]-hydrogenase. We designed broadly inclusive primers targeting the large subunit gene (hhyL) of group 1h [NiFe]-hydrogenases for long-read sequencing to explore its taxonomic distribution across soils. This approach revealed a diverse collection of microorganisms harboring hhyL, including previously unknown groups and taxonomically not assignable sequences. Acidobacterial group 1h [NiFe]-hydrogenase genes were abundant and expressed in temperate soils. To support the participation of acidobacteria in H2 consumption, we studied two representative mesophilic soil acidobacteria, which expressed group 1h [NiFe]-hydrogenases and consumed atmospheric H2 during carbon starvation. This is the first time mesophilic acidobacteria, which are abundant in ubiquitous temperate soils, have been shown to oxidize H2 down to below atmospheric concentrations. As this physiology allows bacteria to survive periods of carbon starvation, it could explain the success of soil acidobacteria. With our long-read sequencing approach of group 1h [NiFe]-hydrogenase genes, we show that the ability to oxidize atmospheric levels of H2 is more widely distributed among soil bacteria than previously recognized and could represent a common mechanism enabling bacteria to persist during periods of carbon deprivation.


Asunto(s)
Acidobacteria , Hidrogenasas , Acidobacteria/metabolismo , Hidrógeno , Hidrogenasas/genética , Hidrogenasas/metabolismo , Oxidación-Reducción , Suelo , Microbiología del Suelo
19.
Appl Environ Microbiol ; 87(3)2021 01 15.
Artículo en Inglés | MEDLINE | ID: mdl-33257309

RESUMEN

Methanogenic archaea have long been implicated in microbially influenced corrosion (MIC) of oil and gas infrastructure, yet a first understanding of the underlying molecular mechanisms has only recently emerged. We surveyed pipeline-associated microbiomes from geographically distinct oil field facilities and found methanogens to account for 0.2 to 9.3% of the 16S rRNA gene sequencing reads. Neither the type nor the abundance of the detected methanogens was correlated with the perceived severity of MIC in these pipelines. Using fluids from one pipeline, MIC was reproduced in the laboratory, both under stagnant conditions and in customized corrosion reactors simulating pipeline flow. High corrosion rates (up to 2.43 mm Fe0 · yr-1) with macroscopic, localized corrosion features were attributed to lithotrophic, mesophilic microbial activity. Other laboratory tests with the same waters yielded negligible corrosion rates (<0.08 mm Fe0 · yr-1). Recently, a novel [NiFe] hydrogenase from Methanococcus maripaludis strain OS7 was demonstrated to accelerate corrosion. We developed a specific quantitative PCR (qPCR) assay and detected the gene encoding the large subunit of this hydrogenase (labeled micH) in corrosive (>0.15 mm Fe0 · yr-1) biofilms. The micH gene, on the other hand, was absent in noncorrosive biofilms, despite an abundance of methanogens. Reconstruction of a nearly complete Methanococcus maripaludis genome from a highly corrosive mixed biofilm revealed micH and associated genes in nearly identical genetic configuration to that in strain OS7, thereby supporting our hypothesis that the encoded molecular mechanism contributed to corrosion. Lastly, the proposed MIC biomarker was detected in multiple oil fields, indicating a geographically widespread involvement of this [NiFe] hydrogenase in MIC.IMPORTANCE Microorganisms can deteriorate built environments, which is particularly problematic in the case of pipelines transporting hydrocarbons to industrial end users. MIC is notoriously difficult to detect and monitor and, as a consequence, is a particularly difficult corrosion mechanism to manage. Despite the advent of molecular tools and improved microbial monitoring strategies for oil and gas operations, specific underlying MIC mechanisms in pipelines remain largely enigmatic. Emerging mechanistic understanding of methanogenic MIC derived from pure culture work allowed us to develop a qPCR assay that distinguishes technically problematic from benign methanogens in a West African oil field. Detection of the same gene in geographically diverse samples from North America hints at the widespread applicability of this assay. The research presented here offers a step toward a mechanistic understanding of biocorrosion in oil fields and introduces a binary marker for (methanogenic) MIC that can find application in corrosion management programs in industrial settings.


Asunto(s)
Proteínas Arqueales/química , Hidrogenasas/química , Residuos Industriales , Yacimiento de Petróleo y Gas , Acero/química , Aguas Residuales/microbiología , Archaea/genética , Archaea/metabolismo , Proteínas Arqueales/genética , Proteínas Arqueales/metabolismo , Carbono , Corrosión , Hidrogenasas/genética , Hidrogenasas/metabolismo , Metano/metabolismo , Reacción en Cadena de la Polimerasa , ARN Ribosómico 16S/genética
20.
Chem Soc Rev ; 50(3): 1668-1784, 2021 Feb 15.
Artículo en Inglés | MEDLINE | ID: mdl-33305760

RESUMEN

While hydrogen plays an ever-increasing role in modern society, nature has utilized hydrogen since a very long time as an energy carrier and storage molecule. Among the enzymatic systems that metabolise hydrogen, [FeFe]-hydrogenases are one of the most powerful systems to perform this conversion. In this light, we will herein present an overview on developments in [FeFe]-hydrogenase research with a strong focus on synthetic mimics and their application within the native enzymatic environment. This review spans from the biological assembly of the natural enzyme and the highly controversial discussed mechanism for the hydrogen generation to the synthesis of multiple mimic platforms as well as their electrochemical behaviour.


Asunto(s)
Materiales Biomiméticos/metabolismo , Hidrogenasas/metabolismo , Materiales Biomiméticos/química , Catálisis , Clostridium/enzimología , Complejos de Coordinación/química , Complejos de Coordinación/metabolismo , Hidrógeno/química , Hidrógeno/metabolismo , Hidrogenasas/química , Hidrogenasas/genética , Metales/química , Mutagénesis Sitio-Dirigida
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA
...