Involvement of a single periplasmic hydrogenase for both hydrogen uptake and production in some Desulfovibrio species

Au cours de cette étude, nous avons montré que plusieurs bactéries sulfato-réductrices possédant un nombre différent de gènes codant pour des hydrogénases, oxydent le lactate en absence de sulfate lorsqu'elles sont en coculture avec #Methanospirillum hungatei. L'efficacité du transfert d'hydrogène avec la bactérie méthanogène n'est pas corrélée avec le nombre de gènes codant pour l'hydrogénase chez ces bactéries sulfato-réductrices. #Desulfovibrio vulgaris Groningen, qui possède uniquement le gène de l'hydrogénase à nickel-fer (hydrogénase [NiFe]), oxyde l'hydrogène en présence de sulfate et produit de l'hydrogène au cours de la fermentation du pyruvate. L'hydrogénase de #D. vulgaris Groningen a été purifiée et caractérisée. Son poids moléculaire est de 87 kDA et elle est constituée de deux sous-unités différentes (60 et 28 kDa). L'hydrogénase de cette bactérie contient 10,6 atomes de fer, 0,9 atome de nickel et 12 atomes de soufre par molécule et son spectre d'absorption est caractéristique d'une protéine à centre fer-soufre. Les activités catalytiques de consommation et production d'hydrogène sont de 332 et 230 unités/mg de protéine, respectivement. Les cellules de #D. vulgarie Groningen contiennent exclusivement l'hydrogénase [NiFe] quelles que soient les conditions de croissance, ainsi que l'ont montré des études biochimiques et immunologiques. L'immunocytolocalisation de cryosections ultrafines de cellules ayant poussé sur différents milieux indique que l'hydrogénase [NiFe] est localisée dans l'espace périplasmique, le marquage étant plus important sur les cellules cultivées sur H2 et sulfate ou pyruvate seul que sur celles cultivées sur lactate et sulfate. Les résultats nous permettent de conclure que #D. vulgaris Groningen contient une seule hydrogénase de type [NiFe] située dans l'espace périplasmique tel que cela a été décrit chez #D. gigas. (Résumé d'auteur

Characterization of Desulfovibrio fructosovorans sp. nov.

Desulfovibrio strain JJ isolated from estuarine sediment differed from all other described Desulfovibrio species by the ability to degrade fructose. The oxidation was incomplete, leading to acetate production. Fructose, malate and fumarate were fermented mainly to succinate and acetate in the absence of an external electron acceptor. The pH and temperature optima for growth were 7.0 and 35° C respectively. Strain JJ was motile by means of a single polar flagellum. The DNA base composition was 64.13% G+C. Cytochrome c3 and desulfoviridin were present. These characteristics established the isolate as a new species of the genus Desulfovibrio, and the name Desulfovibrio fructosovorans is proposed

Wiring of Photosystem II to Hydrogenase for Photoelectrochemical Water Splitting.

In natural photosynthesis, light is used for the production of chemical energy carriers to fuel biological activity. The re-engineering of natural photosynthetic pathways can provide inspiration for sustainable fuel production and insights for understanding the process itself. Here, we employ a semiartificial approach to study photobiological water splitting via a pathway unavailable to nature: the direct coupling of the water oxidation enzyme, photosystem II, to the H2 evolving enzyme, hydrogenase. Essential to this approach is the integration of the isolated enzymes into the artificial circuit of a photoelectrochemical cell. We therefore developed a tailor-made hierarchically structured indium-tin oxide electrode that gives rise to the excellent integration of both photosystem II and hydrogenase for performing the anodic and cathodic half-reactions, respectively. When connected together with the aid of an applied bias, the semiartificial cell demonstrated quantitative electron flow from photosystem II to the hydrogenase with the production of H2 and O2 being in the expected two-to-one ratio and a light-to-hydrogen conversion efficiency of 5.4% under low-intensity red-light irradiation. We thereby demonstrate efficient light-driven water splitting using a pathway inaccessible to biology and report on a widely applicable in vitro platform for the controlled coupling of enzymatic redox processes to meaningfully study photocatalytic reactions.This work was supported by the U.K. Engineering and Physical Sciences Research Council (EP/H00338X/2 to E.R. and EP/G037221/1, nanoDTC, to D.M.), the UK Biology and Biotechnological Sciences Research Council (BB/K002627/1 to A.W.R. and BB/K010220/1 to E.R.), a Marie Curie Intra-European Fellowship (PIEF-GA-2013-625034 to C.Y.L), a Marie Curie International Incoming Fellowship (PIIF-GA-2012-328085 RPSII to J.J.Z) and the CEA and the CNRS (to J.C.F.C.). A.W.R. holds a Wolfson Merit Award from the Royal Society.This is the final version of the article. It first appeared from ACS Publications via http://dx.doi.org/10.1021/jacs.5b0373

The exchange activities of [Fe] hydrogenase (iron–sulfur-cluster-free hydrogenase) from methanogenic archaea in comparison with the exchange activities of [FeFe] and [NiFe] hydrogenases

[Fe] hydrogenase (iron–sulfur-cluster-free hydrogenase) catalyzes the reversible reduction of methenyltetrahydromethanopterin (methenyl-H4MPT+) with H2 to methylene-H4MPT, a reaction involved in methanogenesis from H2 and CO2 in many methanogenic archaea. The enzyme harbors an iron-containing cofactor, in which a low-spin iron is complexed by a pyridone, two CO and a cysteine sulfur. [Fe] hydrogenase is thus similar to [NiFe] and [FeFe] hydrogenases, in which a low-spin iron carbonyl complex, albeit in a dinuclear metal center, is also involved in H2 activation. Like the [NiFe] and [FeFe] hydrogenases, [Fe] hydrogenase catalyzes an active exchange of H2 with protons of water; however, this activity is dependent on the presence of the hydride-accepting methenyl-H4MPT+. In its absence the exchange activity is only 0.01% of that in its presence. The residual activity has been attributed to the presence of traces of methenyl-H4MPT+ in the enzyme preparations, but it could also reflect a weak binding of H2 to the iron in the absence of methenyl-H4MPT+. To test this we reinvestigated the exchange activity with [Fe] hydrogenase reconstituted from apoprotein heterologously produced in Escherichia coli and highly purified iron-containing cofactor and found that in the absence of added methenyl-H4MPT+ the exchange activity was below the detection limit of the tritium method employed (0.1 nmol min−1 mg−1). The finding reiterates that for H2 activation by [Fe] hydrogenase the presence of the hydride-accepting methenyl-H4MPT+ is essentially required. This differentiates [Fe] hydrogenase from [FeFe] and [NiFe] hydrogenases, which actively catalyze H2/H2O exchange in the absence of exogenous electron acceptors