47 research outputs found

    The cytoplasmic domain of the AAA+ protease FtsH is tilted with respect to the membrane to facilitate substrate entry

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    AAA+ proteases are degradation machines that use ATP hydrolysis to unfold protein substrates and translocate them through a central pore toward a degradation chamber. FtsH, a bacterial membrane-anchored AAA+ protease, plays a vital role in membrane protein quality control. How substrates reach the FtsH central pore is an open key question that is not resolved by the available atomic structures of cytoplasmic and periplasmic domains. In this work, we used both negative stain TEM and cryo-EM to determine 3D maps of the full-length Aquifex aeolicus FtsH protease. Unexpectedly, we observed that detergent solubilization induces the formation of fully active FtsH dodecamers, which consist of two FtsH hexamers in a single detergent micelle. The striking tilted conformation of the cytosolic domain in the FtsH dodecamer visualized by negative stain TEM suggests a lateral substrate entrance between the membrane and cytosolic domain. Such a substrate path was then resolved in the cryo-EM structure of the FtsH hexamer. By mapping the available structural information and structure predictions for the transmembrane helices to the amino acid sequence we identified a linker of ∼20 residues between the second transmembrane helix and the cytosolic domain. This unique polypeptide appears to be highly flexible and turned out to be essential for proper functioning of FtsH as its deletion fully eliminated the proteolytic activity of FtsH

    Observation of a new boson at a mass of 125 GeV with the CMS experiment at the LHC

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    Membrane contactor for BaSO4 precipitation : Numerical simulation and experimental results

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    This thesis deals with the study of liquid/liquid tubular membrane contactors. The principle of this process is the following: component A flows through the membrane device inlet to mix/react with component B which comes from the membrane pores. The development of a numerical model showed that mixing between A and B was obtained by diffusion. The use of smaller membrane diameter increased the efficiency of the mixing process. As a consequence, hollow fiber membrane devices were further used for the experimental investigation. The BaSO4 precipitation was chosen as a model reaction. The BaSO4 crystals were analyzed for their size and morphology. Moreover,the conversion rate was measured for several process parameters.Les recherches que nous présentons dans ce mémoire portent sur les contacteurs membranaires en phases liquide/liquide appliqués à la précipitation. Le principe du contacteur à membrane, pour une réaction chimique, est le suivant : un réactif A circule tangentiellement à la surface de la membrane et un réactif B est introduit à travers les pores de la membrane. Le mélange et la réaction entre A et B se produisent à l'intérieur du module membranaire.Un modèle numérique a été développé pour simuler l'écoulement et l'évolution des concentrations des réactifs. Les simulations montrent que l'utilisation de fibres creuses de faible rayon interne permet d'optimiser le mélange. A partir de ces résultats, un pilote a été conçu utilisant des modules fibres creuses pour une réaction modèle : la précipitation de BaSO4 à partir d'une solution de BaCl2 (A) et de K2SO4 (B). L'analyse d'images des cristaux obtenus permet de déterminer leur morphologie, leur taille et le taux de conversion du produit

    Contacteurs à Membrane pour la Précipitation du BaSO4 (simulations numériques et résultats expérimentaux)

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    This thesis deals with the study of liquid/liquid tubular membrane contactors. The principle of this process is the following: component A flows through the membrane device inlet to mix/react with component B which comes from the membrane pores. The development of a numerical model showed that mixing between A and B was obtained by diffusion. The use of smaller membrane diameter increased the efficiency of the mixing process. As a consequence, hollow fiber membrane devices were further used for the experimental investigation. The BaSO4 precipitation was chosen as a model reaction. The BaSO4 crystals were analyzed for their size and morphology. Moreover,the conversion rate was measured for several process parametersLes recherches que nous présentons dans ce mémoire portent sur les contacteurs membranaires en phases liquide/liquide appliqués à la précipitation. Le principe du contacteur à membrane, pour une réaction chimique, est le suivant : un réactif A circule tangentiellement à la surface de la membrane et un réactif B est introduit à travers les pores de la membrane. Le mélange et la réaction entre A et B se produisent à l'intérieur du module membranaire. Un modèle numérique a été développé pour simuler l'écoulement et l'évolution des concentrations des réactifs. Les simulations montrent que l'utilisation de fibres creuses de faible rayon interne permet d'optimiser le mélange. A partir de ces résultats, un pilote a été conçu utilisant des modules fibres creuses pour une réaction modèle : la précipitation de BaSO4 à partir d'une solution de BaCl2 (A) et de K2SO4 (B). L'analyse d'images des cristaux obtenus permet de déterminer leur morphologie, leur taille et le taux de conversion du produitLYON1-BU.Sciences (692662101) / SudocSudocFranceF

    Involvement of mitochondrial ferredoxin and para-aminobenzoic acid in yeast coenzyme Q biosynthesis.: pABA is a precursor of yeast coenzyme Q

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    International audienceYeast ubiquinone or coenzyme Q(6) (Q(6)) is a redox active lipid that plays a crucial role in the mitochondrial electron transport chain. At least nine proteins (Coq1p-9p) participate in Q(6) biosynthesis from 4-hydroxybenzoate (4-HB). We now show that the mitochondrial ferredoxin Yah1p and the ferredoxin reductase Arh1p are required for Q(6) biosynthesis, probably for the first hydroxylation of the pathway. Conditional Gal-YAH1 and Gal-ARH1 mutants accumulate 3-hexaprenyl-4-hydroxyphenol and 3-hexaprenyl-4-aminophenol. Para-aminobenzoic acid (pABA) is shown to be the precursor of 3-hexaprenyl-4-aminophenol and to compete with 4-HB for the prenylation reaction catalyzed by Coq2p. Yeast cells convert U-((13)C)-pABA into (13)C ring-labeled Q(6), a result that identifies pABA as a new precursor of Q(6) and implies an additional NH(2)-to-OH conversion in Q(6) biosynthesis. Our study identifies pABA, Yah1p, and Arh1p as three actors in Q(6) biosynthesis

    Atomistic model of two commercial reverse osmosis membranes

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    Although the main application of reverse osmosis (RO) filtration remains the separation or concentration of electrolytes from aqueous solvents (e.g. desalination), new promising applications including the recycling or purification of organic-rich effluents used in fermentation processes (bio-ethanol and other biofuel productions). Currently, no model exists to predict a priori the selectivity of given RO (mostly in aromatic polyamide) membrane to non-electrolyte organic compounds, such as non-dissociated acids, aldehydes, esters or aromatic compounds. The complication arises due to the mutual diffusion of water and small solutes within membrane, whose swelling is controlled by the stiffness of polymer segments and their cross-linking rates. The general objective of this study is to build atomistic-scale models of typical commercial aromatic polyamide (APA) membranes in order to analyze the contribution of their polymer chemical structure on swelling rate to water and on mutual diffusion mechanisms in the bulk. As APA membranes are polymerized in-situ as approximately 200 nm thick layer, our computational effort was combined with experimental isolation and deformulation of two commercial APA membranes (references: ESPA2 and CPA2, Hydranautics Membranes, USA) to provide both i) initial assumptions to build the atomistic model (monomers composition, average cross-linking rate, approximate swelling rate) and ii) independent reference data to validate the generated model at different relative humidity (X-ray structure factors).The active APA layer was separated from support by removing iteratively each support layer with N,N-dimethylformamide (DMF). Macroscopic properties such as swelling rates, sorption isotherms were assessed by ellipsometry and Intelligent Gravimetric Analyser respectively. A maximum swelling rate of 38% was determined. A cross-linking rate of 65 % was inferred from chemical composition analyses in X-ray photoelectron spectroscopy (XPS) and attenuated-total reflection mode Fourier transform IR (ATR-FTIR). A first atomistic model including the number of desired monomers and water content was built by a conventional compression box technique and classical molecular dynamics code (Discover, Accelrys, San-Diego). Cross-linking was subsequently performed by applying a specific reactive force-field within our own software. The degree of freedoms were i) the initial ratios in monomers (benzene-1,3,5-tricarbonyle chloride with a connectivity of 3, benzene-1,3-diamine with a connectivity of 2), ii) the initial length of block-oligomers (ranging from 5 to 62). Each system was finally equilibrated during long-term isobaric and isothermal molecular dynamics simulations at 600 K and 298 K. Final structure factors were in good agreement with RX scattering spectra

    Atomistic model of two commercial reverse osmosis membranes

    No full text
    Although the main application of reverse osmosis (RO) filtration remains the separation or concentration of electrolytes from aqueous solvents (e.g. desalination), new promising applications including the recycling or purification of organic-rich effluents used in fermentation processes (bio-ethanol and other biofuel productions). Currently, no model exists to predict a priori the selectivity of given RO (mostly in aromatic polyamide) membrane to non-electrolyte organic compounds, such as non-dissociated acids, aldehydes, esters or aromatic compounds. The complication arises due to the mutual diffusion of water and small solutes within membrane, whose swelling is controlled by the stiffness of polymer segments and their cross-linking rates. The general objective of this study is to build atomistic-scale models of typical commercial aromatic polyamide (APA) membranes in order to analyze the contribution of their polymer chemical structure on swelling rate to water and on mutual diffusion mechanisms in the bulk. As APA membranes are polymerized in-situ as approximately 200 nm thick layer, our computational effort was combined with experimental isolation and deformulation of two commercial APA membranes (references: ESPA2 and CPA2, Hydranautics Membranes, USA) to provide both i) initial assumptions to build the atomistic model (monomers composition, average cross-linking rate, approximate swelling rate) and ii) independent reference data to validate the generated model at different relative humidity (X-ray structure factors).The active APA layer was separated from support by removing iteratively each support layer with N,N-dimethylformamide (DMF). Macroscopic properties such as swelling rates, sorption isotherms were assessed by ellipsometry and Intelligent Gravimetric Analyser respectively. A maximum swelling rate of 38% was determined. A cross-linking rate of 65 % was inferred from chemical composition analyses in X-ray photoelectron spectroscopy (XPS) and attenuated-total reflection mode Fourier transform IR (ATR-FTIR). A first atomistic model including the number of desired monomers and water content was built by a conventional compression box technique and classical molecular dynamics code (Discover, Accelrys, San-Diego). Cross-linking was subsequently performed by applying a specific reactive force-field within our own software. The degree of freedoms were i) the initial ratios in monomers (benzene-1,3,5-tricarbonyle chloride with a connectivity of 3, benzene-1,3-diamine with a connectivity of 2), ii) the initial length of block-oligomers (ranging from 5 to 62). Each system was finally equilibrated during long-term isobaric and isothermal molecular dynamics simulations at 600 K and 298 K. Final structure factors were in good agreement with RX scattering spectra

    Atomistic models of polyamide thin films used for the recycling of water in agrofood industries by reverse osmosis

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    Distillation of fermented molasses for alcohol and biofuels production uses large volumes of water. Due to environmental concern, the recycling of condensates in the fermentation medium is encouraged subsequently to a purification step. This step could be performed via reverse osmosis (RO) process, in order to remove in the retentate small organic solutes which act as fermentation inhibitors. Currently, no general rule exists to predict the selectivity of RO membranes according to the nature of organic solutes. Similarly, such insights are missing in nanofiltration processes intended to detoxify biomass hydrolysates prior fermentation. Our objective was to build and validate a molecular model of typical reverse osmosis membranes (exemples: ESPA2 and CPA2 from Hydranautics Membranes, USA), consisting in an active aromatic polyamide layer (APA) with different crosslinking rates. These models will be used in future works to develop general models of transport properties (water swelling, solute partitioning mainly) in RO membranes. The main characteristics of commercial RO membranes were retrieved from isolated APA layers (i.e. separated from the polysulfone-polyester support). Geometrical characteristics such as thickness (about 200 nm), homogeneity and swelling rates were assessed with laser ellipsometry and AFM. The composition and the reticulation rate were inferred from FTIR and XPS measurements. The sorption isotherm of water was sampled with an Intelligent Gravimetric Analyzer. The construction strategy of corresponding atomistic models of dried APA membranes via Molecular Dynamics Simulation (Discover, Accelrys, USA) is presented. The described model gives the theoretical density and neutron scattering spectra of simulated APA. These results could be compared in future works with experimental investigation
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