Crystal Structure of Saccharomyces cerevisiae ECM4, a Xi-Class Glutathione Transferase that Reacts with Glutathionyl-(hydro)quinones

International audienceGlutathionyl-hydroquinone reductases (GHRs) belong to the recently characterized Xi-class of glutathione transferases (GSTXs) according to unique structural properties and are present in all but animal kingdoms. The GHR ScECM4 from the yeast Saccharomyces cerevisiae has been studied since 1997 when it was found to be potentially involved in cell-wall biosyn-thesis. Up to now and in spite of biological studies made on this enzyme, its physiological role remains challenging. The work here reports its crystallographic study. In addition to exhibiting the general GSTX structural features, ScECM4 shows extensions including a huge loop which contributes to the quaternary assembly. These structural extensions are probably specific to Saccharomycetaceae. Soaking of ScECM4 crystals with GS-menadione results in a structure where glutathione forms a mixed disulfide bond with the cysteine 46. Solution studies confirm that ScECM4 has reductase activity for GS-menadione in presence of glutathione. Moreover, the high resolution structures allowed us to propose new roles of conserved residues of the active site to assist the cysteine 46 during the catalytic act

The Warburg Effect Is Genetically Determined in Inherited Pheochromocytomas

The Warburg effect describes how cancer cells down-regulate their aerobic respiration and preferentially use glycolysis to generate energy. To evaluate the link between hypoxia and Warburg effect, we studied mitochondrial electron transport, angiogenesis and glycolysis in pheochromocytomas induced by germ-line mutations in VHL, RET, NF1 and SDH genes. SDH and VHL gene mutations have been shown to lead to the activation of hypoxic response, even in normoxic conditions, a process now referred to as pseudohypoxia. We observed a decrease in electron transport protein expression and activity, associated with increased angiogenesis in SDH- and VHL-related, pseudohypoxic tumors, while stimulation of glycolysis was solely observed in VHL tumors. Moreover, microarray analyses revealed that expression of genes involved in these metabolic pathways is an efficient tool for classification of pheochromocytomas in accordance with the predisposition gene mutated. Our data suggest an unexpected association between pseudohypoxia and loss of p53, which leads to a distinct Warburg effect in VHL-related pheochromocytomas

Impact of early enteral versus parenteral nutrition on mortality in patients requiring mechanical ventilation and catecholamines: study protocol for a randomized controlled trial (NUTRIREA-2)

BACKGROUND: Nutritional support is crucial to the management of patients receiving invasive mechanical ventilation (IMV) and the most commonly prescribed treatment in intensive care units (ICUs). International guidelines consistently indicate that enteral nutrition (EN) should be preferred over parenteral nutrition (PN) whenever possible and started as early as possible. However, no adequately designed study has evaluated whether a specific nutritional modality is associated with decreased mortality. The primary goal of this trial is to assess the hypothesis that early first-line EN, as compared to early first-line PN, decreases day 28 all-cause mortality in patients receiving IMV and vasoactive drugs for shock. METHODS/DESIGN: The NUTRIREA-2 study is a multicenter, open-label, parallel-group, randomized controlled trial comparing early PN versus early EN in critically ill patients requiring IMV for an expected duration of at least 48 hours, combined with vasoactive drugs, for shock. Patients will be allocated at random to first-line PN for at least 72 hours or to first-line EN. In both groups, nutritional support will be started within 24 hours after IMV initiation. Calorie targets will be 20 to 25 kcal/kg/day during the first week, then 25 to 30 kcal/kg/day thereafter. Patients receiving PN may be switched to EN after at least 72 hours in the event of shock resolution (no vasoactive drugs for 24 consecutive hours and arterial lactic acid level below 2 mmol/L). On day 7, all patients receiving PN and having no contraindications to EN will be switched to EN. In both groups, supplemental PN may be added to EN after day 7 in patients with persistent intolerance to EN and inadequate calorie intake. We plan to recruit 2,854 patients at 44 participating ICUs. DISCUSSION: The NUTRIREA-2 study is the first large randomized controlled trial designed to assess the hypothesis that early EN improves survival compared to early PN in ICU patients. Enrollment started on 22 March 2013 and is expected to end in November 2015. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT01802099 (registered 27 February 2013)

Hémophilie A féminine (diagnostic clinique et biologique d'un nouveau cas)

PARIS-BIUP (751062107) / SudocSudocFranceF

La formation en gérontopsychiatrie : effets sur la motivation à travailler auprès de personnes âgées ?

International audienc

Étude cristallographique de glutathion transférases de micro-organismes impliqués dans la dégradation de la lignine (le basidiomycète Phanerochaete chrysosporium et la bactérie Sphingobium sp. SYK-6)

L'étude des champignons saprophytes tel que Phanerochaete chrysosporium est fondamentale car ces organismes, capables de dégrader la lignine, ont un fort potentiel en biotechnologie. Il a été démontré que la bactérie Sphingobium sp. SYK-6 possède des enzymes (ligE, ligF et ligG) appartenant à la famille des glutathion transférases (GST) qui successivement réduisent le lien éther (rôle éthérase) entre les unités de la lignine et déglutathionylent le produit dérivé conjugué (rôle lyase). Ce travail expose les relations entre la structure et la fonction des LigE, F et G de Sphingobium sp. SYK-6 et de deux classes de GST du champignon saprophyte Phanerochaete chrysosporium : une potentiellement liée aux éthérases (GSTFuA) et une potentiellement reliée aux lyases (GST Xi). Les structures cristallographiques des GSTFuA1, 2 et 3 de P. chrysosporium ont été résolues. L'analyse des modèles a permis de révéler une nouvelle classe structurale de GST avec des propriétés uniques. Ces caractéristiques ont pu être reliées à la fonction de ligandine et au profil catalytique de cette nouvelle classe de GST. Parallèlement, les études structurales des LigE et F de Sphingobium sp. SYK-6 sont en voie d'achèvement. La structure cristallographique de la GSTX1 de P. chrysosporium présente des caractéristiques structurales spécifiques qui nous ont conduit à la proposer comme leader d'une nouvelle classe structurale nommée Xi. L'enzyme ne présente pas d'activité lyase vis-à-vis des substrats des LigG, mais en revanche possède une fonction hydroquinone réductase (GHR). Parallèlement, la LigG a été étudiée, et s'apparente structuralement et fonctionnellement la classe Oméga des GSTPhanerochaete chrysosporium is a very interesting saprophytic fungus because it decays wood, by degrading lignin while leaving cellulose which is a renewable source of energy. The soil bacterium Sphingobium sp. SYK-6 possesses enzymes (LigE, LigF and LigG) that cleave the beta-aryl ether linkage of lignin model compounds. LigE and LigF are glutathione dependent enzymes that reduce the ether bond (etherase activity) and LigG catalyzes the elimination of glutathione (lyase activity). This study presents the structure-function relationships of Lig enzymes and of two new classes of GST from P. chrysosporium : one potentially related to LigE (GSTFuA) and one potentially connected to LigG (GST Xi). The crystallographic structures of GSTFuA1, 2 and 3 from P. chrysosporium were solved. The analysis of the models reveals a new structural class of GST with unique properties. These characteristics could be connected to the ligandin function and to the catalytic pattern of this new class of GST. In parallel, structural studies of LigE and LigF from Sphingobium sp. SYK-6 are nearly completed. The crystallographic structure of the GSTX1 from P. chrysosporium exhibits specific structural properties which allowed us to define a new structural class (Xi) in the GST superfamily. The enzyme is a S-glutathionyl-(chloro)hydroquinone reductase (GHR) that does not present a lyase activity with LigG substrate. In parallel, high resolution structure of LigG was obtained; this enzyme can be related structurally and functionally to the GST Omega classMETZ-SCD (574632105) / SudocNANCY1-Bib. numérique (543959902) / SudocNANCY2-Bibliotheque electronique (543959901) / SudocNANCY-INPL-Bib. électronique (545479901) / SudocSudocFranceF

Les logiques implicites de la prestation compensatoire dans le divorce : approches comparées européennes

A une époque où les relations entre les hommes et les femmes sont gouvernées par des principes d’égalité et de liberté, on peut s’interroger sur la pertinence et la justification du maintien d’un droit à une prestation entre les conjoints au lendemain de leur divorce. Les législations européennes s’avérant peu disertes à cet égard, la recherche de la justification du droit à une prestation compensatoire des disparités pécuniaires nécessite l’élaboration préalable d’un outil qui permette de la mettre en évidence. Après avoir éprouvé un modèle diptyque, opposant les traditionnelles natures alimentaire et indemnitaire de la prestation financière après divorce et constaté que ce modèle ne permettait pas de comparaison pertinente entre les droits européens sélectionnés, un second modèle, conçu de manière triptyque, a été élaboré et appliqué à ces différents droits. Ce second modèle dissocie les justifications alimentaire, indemnitaire et assurantielle et permet d’identifier des différences de justification plus marquées, ce qui ne signifie cependant pas pour autant des différences notables entre les différentes prestations après divorce quant aux montants alloués.In an era when relations between men and women are governed by the principles of equality and liberty, we may ask whether it is appropriate to preserve a post-divorce support entitlement between spouses, and what justification there is for doing so. European legislation is not very forthcoming in this regard, and this means that looking for the justification for entitlement to an allowance to compensate for financial disparities calls for a tool to be developed first that would identify that justification. After testing a two-option model in which the traditional support and compensation functions of a post-divorce financial allowance were contrasted and finding that this model did not provide a useful comparison between the selected European legal systems, a second, three-option model was developed and applied to those systems. The second model distinguishes the support, compensation and insurance justifications and can identify more pronounced differences. However, this does not mean that there are any noteworthy differences among the various post-divorce allowances in terms of the amounts awarded

Caractérisation cristallographique d'intermédiaires réactionnels de méthionine sulfoxyde réductases en vue de la compréhension de leur mécanisme catalytique (Les trois domaines de la protéine multifonctionnelle PilB de Neisseria meningitidis et la MsrB de Xanthomonas campestris)

Les résidus méthionine sont facilement oxydables en sulfoxydes in vivo. Cette oxydation est réversible via la méthionine sulfoxyde réductase (Msr), un enzyme ubiquitaire. Dû aux deux configurations possibles du sulfoxyde, deux classes d'enzyme structuralement différents existent : les MsrA sont spécifiques de l'isomère S, et les MsrB de l'isomère R. Les deux possèdent un même mécanisme catalytique en deux étapes. La première étape est dédiée à la réduction du substrat ; elle aboutit à l oxydation d une cystéine en acide sulfénique. Puis le recyclage de l enzyme s opère avec la formation d un pont disulfure intramoléculaire qui est finalement réduit par une thiorédoxine (Trx). Chez Neisseria meningitidis la protéine PilB porte MsrA et MsrB sous forme de deux domaines. Un troisième domaine à activité du type Trx existe en N-terminal. Les trois domaines isolés ont été étudiés par cristallographie. (i) Le domaine N-terminal : la structure confirme son homologie avec la Trx, mais son analyse révèle des éléments probablement à l origine d un fonctionnement particulier. (ii) La MsrA : la structure de deux mutants a permis d observer un complexe avec un substrat et l acide sulfénique. Leurs études s additionnent à celles de l enzyme sauvage sous formes réduite et oxydée. (iii) La MsrB : la structure d un mutant a également permis d obtenir un complexe et complète les structures des formes réduite et oxydée du sauvage. En addition, la structure de la MsrB de Xanthomonas campestris met à jour des différences conformationnelles entre MsrB d organismes différents. Enfin, l étude structurale de PilB entier a été entamée en solution par la méthode de diffusion aux petits angles.Methionine residues are easily oxidized to sulfoxides, in vivo. This oxidation is reversed via a ubiquitous enzyme named methionine sulfoxide reductase (Msr). Due to the two possible configurations of the sulfoxide group, two structurally-different classes of enzymes exist: MsrAs are specific for the isomer S, and MsrBs for the isomer R. Both classes act through a two-step mechanism. The first step is dedicated to substrate reduction. It results in the sulfenic form of the catalytic cysteine. Then recycling of the enzyme starts with the formation of an intramolecular disulfide bridge, finally reduced by thioredoxin (Trx). In Neisseria meningitidis, the protein PilB bears MsrA and MsrB as two adjacent domains. A third domain with a Trx-like activity exists at the N-terminal end. The three isolated domains have been studied by X-Ray crystallography. (i) The N-terminal domain: its structure confirmed its homology to Trx and DsbEs, but its analysis revealed elements that probably explain its peculiar properties. (ii) MsrA: the structures of two mutants allowed the observation of a complex with the substrate and of the sulfenic acid. These results add to the structure of the reduced and oxidized forms of the wild type domain so that the catalytic mechanism can be analyzed. (iii) MsrB: the structures of the reduced and oxidized forms were completed by that of a complex with the substrate obtained from a mutant. In addition, comparison with the Xanthomonas campestris MsrB structure enlightened conformational differences between MsrBs from distinct organisms. Finally, structural studies of the whole PilB protein have been initiated in solution using small angle X-Ray scattering.NANCY1-Bib. numérique (543959902) / SudocSudocFranceF

Seuils d'âge en droit des personnes et de la famille

Recherche JEUVUL ou Droit et Jeunesse. Recherche, labélisée et financée par la Maison des sciences de l'homme de Bretagne (MSHB)International audienc

Disulphide dimerised peptide creates a crystal contact in an anti-peptide antibody

International audienc