Shewanella oneidensis: a new and efficient System for Expression and Maturation of heterologous [Fe-Fe] Hydrogenase from Chlamydomonas reinhardtii

<p>Abstract</p> <p>Background</p> <p>The eukaryotic green alga, <it>Chlamydomonas reinhardtii</it>, produces H₂ under anaerobic conditions, in a reaction catalysed by a [Fe-Fe] hydrogenase HydA1. For further biochemical and biophysical studies a suitable expression system of this enzyme should be found to overcome its weak expression in the host organism. Two heterologous expression systems used up to now have several advantages. However they are not free from some drawbacks. In this work we use bacterium <it>Shewanella oneidensis</it> as a new and efficient system for expression and maturation of HydA1 from <it>Chlamydomonas reinhardtii</it>.</p> <p>Results</p> <p>Based on codon usage bias and hydrogenase maturation ability, the bacterium <it>S. oneidensis</it>, which possesses putative [Fe-Fe] and [Ni-Fe] hydrogenase operons, was selected as the best potential host for <it>C. reinhardtii </it>[Fe-Fe] hydrogenase expression. Hydrogen formation by <it>S. oneidensis </it>strain AS52 (Δ<it>hydA</it>Δ<it>hyaB</it>) transformed with a plasmid bearing <it>Cr</it>HydA1 and grown in the presence of six different substrates for anaerobic respiration was determined. A significant increase in hydrogen evolution was observed for cells grown in the presence of trimethylamine oxide, dimethylsulfoxide and disodium thiosulfate, showing that the system of <it>S. oneidensis </it>is efficient for heterologous expression of algal [Fe-Fe] hydrogenase.</p> <p>Conclusion</p> <p>In the present work a new efficient system for heterologous expression and maturation of <it>C. reinhardtii </it>hydrogenase has been developed. HydA1 of <it>C. reinhardtii </it>was purified and shown to contain 6 Fe atoms/molecule of protein, as expected. Using DMSO, TMAO or thiosulfate as substrates for anaerobic respiration during the cell growth, 0.4 – 0.5 mg l^-1(OD₆₀₀= 1) of catalytically active HydA1 was obtained with hydrogen evolution rate of ~700 μmol H₂mg^-1min^-1.</p

P2CS: a two-component system resource for prokaryotic signal transduction research

BACKGROUND: With the escalation of high throughput prokaryotic genome sequencing, there is an ever-increasing need for databases that characterise, catalogue and present data relating to particular gene sets and genomes/metagenomes. Two-component system (TCS) signal transduction pathways are the dominant mechanisms by which micro-organisms sense and respond to external as well as internal environmental changes. These systems respond to a wide range of stimuli by triggering diverse physiological adjustments, including alterations in gene expression, enzymatic reactions, or protein-protein interactions. DESCRIPTION: We present P2CS (Prokaryotic 2-Component Systems), an integrated and comprehensive database of TCS signal transduction proteins, which contains a compilation of the TCS genes within 755 completely sequenced prokaryotic genomes and 39 metagenomes. P2CS provides detailed annotation of each TCS gene including family classification, sequence features, functional domains, as well as genomic context visualization. To bypass the generic problem of gene underestimation during genome annotation, we also constituted and searched an ORFeome, which improves the recovery of TCS proteins compared to searches on the equivalent proteomes. CONCLUSION: P2CS has been developed for computational analysis of the modular TCSs of prokaryotic genomes and metagenomes. It provides a complete overview of information on TCSs, including predicted candidate proteins and probable proteins, which need further curation/validation. The database can be browsed and queried with a user-friendly web interface at

The Cyst-Dividing Bacterium Ramlibacter tataouinensis TTB310 Genome Reveals a Well-Stocked Toolbox for Adaptation to a Desert Environment

Ramlibacter tataouinensis TTB310T (strain TTB310), a betaproteobacterium isolated from a semi-arid region of South Tunisia (Tataouine), is characterized by the presence of both spherical and rod-shaped cells in pure culture. Cell division of strain TTB310 occurs by the binary fission of spherical “cyst-like” cells (“cyst-cyst” division). The rod-shaped cells formed at the periphery of a colony (consisting mainly of cysts) are highly motile and colonize a new environment, where they form a new colony by reversion to cyst-like cells. This unique cell cycle of strain TTB310, with desiccation tolerant cyst-like cells capable of division and desiccation sensitive motile rods capable of dissemination, appears to be a novel adaptation for life in a hot and dry desert environment. In order to gain insights into strain TTB310's underlying genetic repertoire and possible mechanisms responsible for its unusual lifestyle, the genome of strain TTB310 was completely sequenced and subsequently annotated. The complete genome consists of a single circular chromosome of 4,070,194 bp with an average G+C content of 70.0%, the highest among the Betaproteobacteria sequenced to date, with total of 3,899 predicted coding sequences covering 92% of the genome. We found that strain TTB310 has developed a highly complex network of two-component systems, which may utilize responses to light and perhaps a rudimentary circadian hourglass to anticipate water availability at the dew time in the middle/end of the desert winter nights and thus direct the growth window to cyclic water availability times. Other interesting features of the strain TTB310 genome that appear to be important for desiccation tolerance, including intermediary metabolism compounds such as trehalose or polyhydroxyalkanoate, and signal transduction pathways, are presented and discussed

Enjeux théoriques de la délocalisation

Comme le rappelle l'introduction du rapport du Conseil d'analyse économique de Fontagné et Lorenzi (2005), le thème des délocalisations s'est récemment imposé dans le débat politique et social en France, et plus largement dans les pays industrialisés. Les délocalisations, que l'on peut définir comme des fermetures d'unités de production dans l'économie nationale suivies de réouvertures à l'étranger, s'inscrivent dans le contexte plus large d'une réorganisation globale des activités des entreprises sur une base mondiale. À ce titre, les délocalisations peuvent être analysées comme de l'investissement direct à l'étranger (IDE). La théorie économique distingue généralement deux types d'IDE. L'IDE dit horizontal est essentiellement motivé par des questions d'accès au marché (« market seeking »). Du point de vue stratégique, il est alors considéré comme le choix d'une entreprise de produire directement sur le marché local plutôt que de produire dans son pays d'origine pour exporter ensuite. L'IDE dit vertical obéit quant à lui à des motifs de recherche d'efficacité (« efficiency seeking »), le choix d'implantation d'unités de production à l'étranger reposant alors sur une comparaison de coûts de production. Il s'inscrit souvent dans une stratégie globale de fragmentation de la production, différents éléments du produit final étant alors produits dans différents pays

Control and Regulation of KplE1 Prophage Site-specific Recombination

International audienc

Comparison of Methods for Quantification of Cytochrome cd(1)-Denitrifying Bacteria in Environmental Marine Samples

Two PCR primer sets were developed for the detection and quantification of cytochrome cd(1)-denitrifying bacteria in environmental marine samples. The specificity and sensitivity of these primers were tested. Both primer sets were suitable for detection, but only one set, cd3F–cd4R, was suitable for the quantification and enumeration of the functional community using most-probable-number PCR and competitive PCR techniques. Quantification of cytochrome cd(1) denitrifiers taken from marine sediment and water samples was achieved using two different molecular techniques which target the nirS gene, and the results were compared to those obtained by using the classical cultivation method. Enumerations using both molecular techniques yielded similar results in seawater and sediment samples. However, both molecular techniques showed 1,000 or 10 times more cytochrome cd(1) denitrifiers in the sediment or water samples, respectively, than were found by use of the conventional cultivation method for counting

Bacterial physiology: Hsp90 is the hot guy for Shewanella oneidensis

International audienc

TorI, a response regulator inhibitor of phage origin in Escherichia coli

The torI gene has been identified by using a genetic multicopy approach as a negative regulator of the torCAD operon that encodes the trimethylamine N-oxide reductase respiratory system in Escherichia coli. The negative effect was due to a previously unidentified small ORF (66 aa) of phage origin that we called torI for Tor inhibition. Overexpression of torI led to an 8-fold decrease of the torCAD operon transcription. This operon is positively regulated, in the presence of trimethylamine N-oxide, by a four-step phosphorelay involving the TorS sensor and the TorR response regulator. Epistatic experiments showed that TorI acts downstream of TorS and needs the presence of TorR. In vitro experiments showed that it is neither a TorR phosphatase nor a histidine kinase inhibitor and that it binds to the effector domain of TorR. Unexpectedly, TorI did not impede TorR DNA binding, and we propose that it may prevent RNA polymerase recruitment to the torC promoter. This study thus reveals a previously uncharacterized class of response regulator inhibitors