Mitochondrial ADP/ATP Carrier in Dodecylphosphocholine Binds Cardiolipins with Non-native Affinity

Biophysical investigation of membrane proteins generally requires their extraction from native sources using detergents, a step that can lead, possibly irreversibly, to protein denaturation. The propensity of docecylphosphocholine (DPC), a detergent widely utilized in NMR studies of membrane proteins, to distort their structure has been the subject of much controversy. It has been recently proposed that the binding specificity of the yeast mitochondrial ADP/ATP carrier (yAAC3) toward cardiolipins (CLs) is preserved in DPC, thereby suggesting that DPC is a suitable environment to study membrane proteins. In this communication, we used all-atom molecular dynamics simulations to investigate the specific binding of CLs to yAAC3. Our data demonstrate that the interaction interface observed in a native-like environment differs markedly from that inferred from an NMR investiga- tion in DPC, implying that in this detergent, the protein structure is distorted. We further investigated yAAC3 solubilized in DPC and in the milder dodecylmaltoside with thermal-shift assays. The loss of thermal transition observed in DPC confirms that the protein is no longer properly folded in this environment

Mechanism of the allosteric activation of the ClpP protease machinery by substrates and active-site inhibitors

Coordinated conformational transitions in oligomeric enzymatic complexes modulate function in response to substrates and play a crucial role in enzyme inhibition and activation. Caseinolytic protease (ClpP) is a tetradecameric complex, which has emerged as a drug target against multiple pathogenic bacteria. Activation of different ClpPs by inhibitors has been independently reported from drug development efforts, but no rationale for inhibitor-induced activation has been hitherto proposed. Using an integrated approach that includes x-ray crystallography, solid- and solution-state nuclear magnetic resonance, molecular dynamics simulations, and isothermal titration calorimetry, we show that the proteasome inhibitor bortezomib binds to the ClpP active-site serine, mimicking a peptide substrate, and induces a concerted allosteric activation of the complex. The bortezomib-activated conformation also exhibits a higher affinity for its cognate unfoldase ClpX. We propose a universal allosteric mechanism, where substrate binding to a single subunit locks ClpP into an active conformation optimized for chaperone association and protein processive degradation

Mechanism of the allosteric activation of the ClpP protease machinery by substrates and active-site inhibitors

18 pags., 6 figs., 1 tab. -- Open Access funded by Creative Commons Atribution Licence 4.0Coordinated conformational transitions in oligomeric enzymatic complexes modulate function in response to substrates and play a crucial role in enzyme inhibition and activation. Caseinolytic protease (ClpP) is a tetradecameric complex, which has emerged as a drug target against multiple pathogenic bacteria. Activation of different ClpPs by inhibitors has been independently reported from drug development efforts, but no rationale for inhibitor-induced activation has been hitherto proposed. Using an integrated approach that includes x-ray crystallography, solid- and solution-state nuclear magnetic resonance, molecular dynamics simulations, and isothermal titration calorimetry, we show that the proteasome inhibitor bortezomib binds to the ClpP active-site serine, mimicking a peptide substrate, and induces a concerted allosteric activation of the complex. The bortezomib-activated conformation also exhibits a higher affinity for its cognate unfoldase ClpX. We propose a universal allosteric mechanism, where substrate binding to a single subunit locks ClpP into an active conformation optimized for chaperone association and protein processive degradation.This work used the platforms of the Grenoble Instruct center (ISBG; UMS 3518 CNRS-CEA-UJF-EMBL) with support from INSTRUCT (“Innovative EM/NMR approach for the characterization of the drug target ClpP APPID: 301“), FRISBI (ANR-10-INSB-05-02), and GRAL (ANR-10-LABX-49-01) within the Grenoble Partnership for Structural Biology (PSB). We thank the ESRF for beamtime at ID30A and ID23-1. Funding: This work was supported by Spanish Ministerio de Economia y Competitividad (BFU2016-78232-P) and Instituto de Salud Carlos III co-funded by European Union (PI15/00663 and PI18/00349, ERDF/ ESF, “Investing in your future”). This work was financially supported by the European Research Council (ERC-Stg-2012-311318 to P.S.). J.F. is supported by an EMBO long-term post-doctoral fellowship (ALTF441-2017)

Intensity Weighted Subtraction Microscopy Approach for Image Contrast and Resolution Enhancement

We propose and demonstrate a novel subtraction microscopy algorithm, exploiting fluorescence emission difference or switching laser mode and their derivatives for image enhancement. The key novelty of the proposed approach lies in the weighted subtraction coefficient, adjusted pixel-by-pixel with respect to the intensity distributions of initial images. This method produces significant resolution enhancement and minimizes image distortions. Our theoretical and experimental studies demonstrate that this approach can be applied to any optical microscopy techniques, including label free and non-linear methods, where common super-resolution techniques cannot be used

Transition énergétique et développement territorial, une promesse à questionner

National audienceCette recherche questionne les promesses et les ambiguïtés de la transition énergétique en s'intéressant aux modalités de déploiement des projets bois-énergie en Aquitaine. Quelles sont les synergies et les valeurs qui sous-tendent leur émergence et leur ancrage aux territoires ? L'enquête menée souligne le poids des héritages territoriaux dans la diversité des « communautés de valeurs » construites, de la défense d'un tissu industriel à la réhabilitation d'un service public de l'énergie. Elle montre aussi que l'approvisionnement local est un levier important mais pas forcément une nécessité, ni un gage de durabilité pour ces territoires. Elle souligne enfin l'inventivité déployée pour s'adapter aux attendus croissants concernant la performance économique et environnementale des équipements installés. Cette réflexion invite in fine à mettre l'accent sur les enjeux de solidarité dans la production des communautés énergétiques durables et à développer des pratiques de planification adaptées à des sources d'énergie présentes partout mais en faible densité et de manière intermittent

Propositions en vue d'une amélioration pour la mesure et le suivi de l'indicateur C6.10 au sein du processus Forest Europe

The primary objective of this report is to assess the current FOREST EUROPE Indicator 6.10 and to make recommendations to improve the reporting of the indicator. The report also considers recreation monitoring to better understand the reasons behind the difficulties in reporting this FOREST EUROPE indicator. The second objective is to identify means to improve monitoring of Indicator 6.10, and assess the use of recreation indicators in selected European countries as a means to identify best practice in terms of a stable standard and quality of measurements, which are comparable across European countries and over time. The report is the result of the work by a small group of experts, mainly researchers, from Finland, Sweden, Denmark, Switzerland, Austria, France and the UK