La dystrophine est une très grande protéine codée le gène DMD et située sous la membrane plasmique des fibres musculaires. Elle joue un rôle essentiel dans le maintien de l’intégrité de la cellule musculaire lors des cycles de contraction/relaxation. Cette protéine filamenteuse est composée de quatre domaines structuraux dont le domaine central composé de 24 répétitions homologues à la spectrine. Chaque répétition est organisée en faisceau de trois α-hélices appelé « coiled-coil ». Des mutations du gène DMD sont à l’origine des myopathies de Duchenne (DMD) et de Becker (BMD) qui s’accompagnent d’un déficit total ou d’une dystrophine mutée et induisent de ruptures fréquentes de la membrane des cellules musculaires. La connaissance de la structure de la dystrophine est nécessaire au développement de thérapies à ce jour inexistantes pour les myopathies. Au laboratoire, des données structurales du domaine central de la dystrophine ont été acquises par diffusion des rayons X aux petits angles (SAXS, Small Angles X-ray Scattering). Cette thèse présente le développement d’une approche multi-échelle combinant des données expérimentales SAXS et des données in silico pour la reconstruction de modèles haute-résolution des fragments du domaine central de la dystrophine et d’un fragment muté observé dans une mutation BMD fréquente. Nous avons également cartographié l’interaction de ce domaine central avec deux de ses partenaires fonctionnels importants, l’actine filamenteuse et avec la nitroxyde synthase neuronale (nNOS) et proposé les premiers modèles atomiques des complexes macromoléculaires correspondants. L’ensemble de ces résultats permettra à terme l’optimisation de thérapies pour le traitement des dystrophies musculaires. / Dystrophin is a large protein encoded by DMD gene and located under the plasma membrane of muscle fibers. It plays an essential role in maintaining the integrity of muscle cells during contraction/relaxation cycles. This filamentous protein is composed of four structural domains including the central domain consisting of 24 spectrin-like repeats and four hinges. Each repetition is folded in three α-helices in a ‘coiled-coil’ assembly. Mutations in the DMD gene leads to Duchenne muscular dystrophy (DMD) and Becker (MDBs), which are accompanied by frequent plasma membrane ruptures, due to the loss or modification of dystrophin protein. There are very few structural data available concerning the central domain of dystrophin, which is subject to many mutations involved in DMD and BMD diseases. However, the description and the understanding to an atomic level of dystrophin structure and its interaction is essential for optimization of therapies. Given the impossibility to solve its structure by X-ray crystallography or NMR, structural data of the dystrophin central domain were acquired by small angles X-rays scattering (SAXS, Small Angles X-ray Scattering). This thesis presents the development of an innovative multi-scale approach combining experimental SAXS and in silico derived data, allowing the reconstruction of high-resolution models of dystrophin central domain fragments. Structural data were also obtained on a mutated dystrophin frequently observed in BMDs. Furthermore, we also mapped the interactions of the central domain with two of its majors functional partners, Filamentous actin and neuronal nitroxyde synthase (nNOS) and proposed models of the related macromolecular complexes. At long-term, all of these results will allow optimization of therapies for the treatment of muscular dystrophies.

Coq6 est une enzyme impliquée dans la biosynthèse du coenzyme Q (aussi nommé ubiquinone, ou Q), un lipide benzoquinone polyprenylé essentiel à la fonction de la chaîne respiratoire mitochondriale. Dans la levure Saccharomyces cerevisiae, cette monooxygénase flavine-dépendante putatif est proposé pour hydroxyler le noyau benzénique d' un précurseur du coenzyme Q à la position C5. Nous montrons ici à travers des études biochimiques que Coq6 est une flavoprotéine utilisant le FAD comme cofacteur. Des modèles d'homologie du complexe Coq6-FAD ont étés réalisés et étudiés par dynamique moléculaire et arrimage moléculaire du 3-hexaprenyl-4-hydroxyphényl (4-HP6), un substrat modèle hydrophobe et volumineux. Nous identifions un canal d'accès putatif pour Coq6 dans un modèle de la forme sauvage et proposons des mutations in silico positionnés à l'entrée capable de partiellement (les mutations simples G248R et L382E) ou complètement (une double-mutation G248R-L382E) bloquer l'accès du substrat au site actif via le canal d' accès. Des essais in vivo soutiennent les prédictions in silico, qui expliquent l'abrogation ou la diminution des enzymes mutées. Ce travail fournit la première information structurale détaillée d'une enzyme importante et hautement conservée de biosynthèse de l'ubiquinone. / Coq6 is an enzyme involved in the biosynthesis of coenzyme Q, a polyisoprenylated benzoquinone lipid essential to the function of the mitochondrial respiratory chain. In the yeast Saccharomyces cerevisiae, this putative flavin-dependent monooxygenase is proposed to hydroxylate the benzene ring of coenzyme Q (ubiquinone) precursor at position C5. We show here through biochemical studies that Coq6 is a flavoprotein using FAD as a cofactor. Homology models of the Coq6-FAD complex are constructed and studied through molecular dynamics and substrate docking calculations of 3-hexaprenyl-4-hydroxyphenol (4-HP6), a bulky hydrophobic model substrate. We identify a putative access channel for Coq6 in a wild type model and propose in silico mutations positioned at its entrance capable of partially (G248R and L382E single mutations) or completely (a G248R-L382E double-mutation) blocking access of the substrate to thechannel . Further in vivo assays support the computational predictions, thus explaining the decreased activities or inactivation of the mutated enzymes. This work provides the first detailed structural information of an important and highly conserved enzyme of ubiquinone biosynthesis.

Chloroperoxidase (CPO) is the most versatile heme-containing enzyme that catalyzes a broad spectrum of reactions. The remarkable feature of this enzyme is the high regio- and enantio-selectivity exhibited in CPO-catalyzed oxidation reactions. The aim of this dissertation is to elucidate the structural basis for regio- and enantio-selective transformations and investigate the application of CPO in biodegradation of synthetic dyes. To unravel the mechanism of CPO-catalyzed regioselective oxidation of indole, the dissertation explored the structure of CPO-indole complex using paramagnetic relaxation and molecular modeling. The distances between the protons of indole and the heme iron revealed that the pyrrole ring of indole is oriented toward the heme with its 2-H pointing directly at the heme iron. This provides the first experimental and theoretical explanation for the "unexpected" regioselectivity of CPO-catalyzed indole oxidation. Furthermore, the residues including Leu 70, Phe 103, Ile 179, Val 182, Glu 183, and Phe 186 were found essential to the substrate binding to CPO. These results will serve as a lighthouse in guiding the design of CPO mutants with tailor-made activities for biotechnological applications. To understand the origin of the enantioselectivity of CPO-catalyzed oxidation reactions, the interactions of CPO with substrates such as 2-(methylthio)thiophene were investigated by nuclear magnetic resonance spectroscopy (NMR) and computational techniques. In particular, the enantioselectivity is partly explained by the binding orientation of substrates. In third facet of this dissertation, a green and efficient system for degradation of synthetic dyes was developed. Several commercial dyes such as orange G were tested in the CPO-H2O2-Cl- system, where degradation of these dyes was found very efficient. The presence of halide ions and acidic pH were found necessary to the decomposition of dyes. Significantly, the results revealed that this degradation of azo dyes involves a ferric hypochlorite intermediate of CPO (Fe-OCl), compound X.

This work explores the use of computational methods to aid in the design of Metal Organic Frameworks (MOFs) for use as CO2 scrubbers in carbon capture and storage applications. One of the main challenges in this field is in identifying important MOF design characteristics which optimize the complex interactions governing surface adsorption. We approach this in a high-throughput manner, determining properties important to CO2 adsorption from generating and sampling a large materials search space. The utilization of MOFs as potential carbon scrubbing agents is a recent phenomenon, as such, many of the computational tools necessary to perform high-throughput screening of MOFs and subsequent analysis are either underdeveloped or non-existent. A large portion of this work therefore involved the development of novel tools designed specifically for this task. The chapters in this work are contiguous with the goal of designing MOFs for CO¬2 capture, and somewhat chronological in order and complexity, meaning as time and expertise progressed, more advanced tools were developed and utilized for the purposes of computational MOF discovery. Initial work towards MOF design involved the detailed analysis of two experimental structures; CALF-15 and CALF-16 using classical molecular dynamics, grand canonical Monte Carlo simulations, and DFT to determine the structural features which promote CO2 adsorption. An unprecedented level of agreement was found between theory and experiment, as we are able to capture, with simulation, the X-ray resolved binding sites of CO2 in the confined pores of CALF-15. Molecular simulation was then used to provide a detailed breakdown of the energy contributions from nearby functional groups in both CALF-15 and CALF-16. A large database of hypothetical MOF structures is constructed for the purposes of screening for CO2 adsorption. The database contains 1.3 million hypothetical structures, generated with an algorithm which snaps together rigid molecular building blocks extracted from existing MOF crystal structures. The algorithm for constructing the hypothetical MOFs and the building blocks themselves were all developed in-house to form the resulting database. The topological, chemical, and physical features of these MOFs are compared to recently developed materials databases to demonstrate the larger structural and chemical space sampled by our database. In order to rapidly and accurately describe the electrostatic interactions of CO2 in the hypothetical database of MOFs, parameters were developed for use with the charge equilibration method. This method assigns partial charges on the framework atoms based on a set of parameters assigned to each atom type. An evolutionary algorithm was used to optimize the charge equilibration parameters on a set of 543 hypothetical MOFs such that the partial charges generated would reproduce each MOFs DFT-derived electrostatic potential. Validation of these parameters was performed by comparing the CO2 adsorption from the charge equilibration method vs DFT-derived charges on a separate set of 693 MOFs. Our parameter set were found to reproduce DFT-derived CO2 adsorption extremely well using only a fraction of the time, making this method ideal for rapid and accurate high-throughput MOF screening. A database of 325,000 MOFs was then screened for CO2 capture and storage applications. From this study we identify important binding pockets for CO2 in MOFs using a binding site analysis tool. This tool uses a pattern recognition method to compare the 3-D configurations of thousands of pore structures surrounding strong CO2 adsorption sites, and present common features found amongst them. For the purposes of developing larger databases which sample a more diverse materials space, a novel MOF construction tool is devloped which builds MOFs based on abstract graphs. The graph theoretical foundations of this method are discussed and several examples of MOF construction are presented to demonstrate its use. Notably, not only can it build existing MOFs with complicated geometries, but it can sample a wide range of unique structures not yet discovered by experimental means.

Traditional computational protein design (CPD) calculations model sequence perturbations and evaluate their stabilities using a single fixed protein backbone template in an approach referred to as single‐state design (SSD). However, certain design objectives require the explicit consideration of multiple conformational states. Cases where a multistate framework may be advantageous over the single‐state approach include the computer aided discovery of new enzyme substrates, the prediction of protein stabilities, and the design of protein dynamics. These design objectives can be tackled using multistate design (MSD). However, it is often the case that a design objective requires the consideration of a protein state having no available structure information. For such circumstances the multistate framework cannot be applied. In this thesis I present the development of two template and ensemble preparation methodologies and their application to three projects. The purpose of which is to demonstrate the necessary ensemble modeling strategies to overcome limitations in available structure information. Particular emphasis is placed on the ability to recapitulate experimental data to guide modelling of the design space. Specifically, the use of MSD allowed for the accurate prediction of a methyltransferase recognition motif and new substrates, the prediction of mutant sequence stabilities with quantitative accuracy, and the design of dynamics into the rigid Gβ1 scaffold producing a set of dynamic variants whose tryptophan residue exchanges between two conformations on the millisecond timescale. Implementation of both the ensemble, coordinate perturbation followed by energy minimization (PertMin), and template, rotamer optimization followed by energy minimization (ROM), generation protocols developed here allow for exploration and manipulation of the structure space enabling the success of these applications.

Le sujet de cette thèse concerne les études structurales d'une nouvelle génération d'oligomères, les foldamères, construits à partir de mimes pseudopeptidiques capables d'adopter des structures secondaires déterminées. Trois familles d'oligomères ont été étudiées. Plusieurs méthodes spectroscopiques ont été à chaque fois employées couplées à la modélisation moléculaire, afin de caractériser les propriétés structurales des foldamères. La première famille a été synthétisée par le groupe de Muriel Amblard et Monique Calmès de l'IBMM à Montpellier. Nous avons évalué les propriétés à se structurer du motif (S)-ABOC. Plusieurs oligomères de tailles variables ont été synthétisés et nous avons montré le motif (S)ABOC lorsqu'il est introduit dans une séquence peptidique est capable de générer des coudes beta. L'enchaînement de ces coudes conduits à la formation d'hélices qui en fonction des motifs pseudo-peptidiques associés présentent des caractéristiques différentes. La seconde famille étudiée comprenait des oligomères formés à partir de monomères contraints de type thiazole. Nous avons montré que ce motif, développé par Ludovic Maillard à l'IBMM à Montpellier forme des pseudocycles en C9 qui s'organisent ensuite en une hélice qui est observable dès le tétramère. La troisième famille d‘oligomères étudiée est issue du LCPM. Elle comprend des oligomères alternant des acides alpha-aminés et des acides aminés de type aza dans lesquels le carbone alpha a été remplacé par un atome d'azote. Nous avons montré que le motif aza induit un repliement en coude beta facilitant la cyclisation de la molécule. Dans ma thèse nous avons caractérisé les propriétés à s'auto-structurer de trois familles de foldamères pseudopeptidiques en combinant les résultats expérimentaux issus de la résonance magnétique nucléaire, la spectroscopie infrarouge, le dichroïsme circulaire et la diffraction des rayons X et en les intégrant à des calculs de modélisation moléculaire. Notre étude répertorie les propriétés de chaque motif à former des éléments de structure secondaire et à induire la formation de foldamères / The subject of this thesis deals with the structural studies of a new generation of oligomers, so called foldamers, constructed from pseudopeptidic mimics and able to adopt determined secondary structures. Three families of oligomers were investigated. For each oligomer several spectroscopic methods have been used combined with molecular modeling in order to characterize their structural properties. The first family was studied in collaration with the group of Muriel Amblard and Monique Calmès at IBMM in Montpellier. We have evaluated the structural properties of the motif (S)-ABOC. Several oligomers of varying sizes were synthesized and we have shown that the pattern (S)-ABOC, when introduced into a peptide sequence, is capable of generating beta turn. The sequence of these turn leads to the formation of helices, which, according to associated pseudo-peptide patterns shows different characteristics. The second project discussed the conformation of oligomers family formed from constrainted thiazole monomers. We have shown that this pattern developed by Ludovic Maillard at IBMM in Montpellier adopted pseudocycles C9 which led to helices. THis property was observed starting from the tetramer. The third family of oligomers comes from the LCPM, alternating oligomers of alpha amino acids and aza amino acids, in which the alpha carbon has been replaced by a nitrogen atom. We showed that the pattern aza induces folding of the beta turn, facilitating the cyclization of the molecule. In this thesis we have characterized the properties to self-structuring of three families of pseudopeptide foldamers by combining experimental results from nuclear magnetic resonance, infrared spectroscopy, circular dichroism and X-ray diffraction combined with molecular modeling calculations. Our study lists the properties of each pattern to form secondary structure elements and to induce the formation of foldamers

The effect of impurities on crystal morphology is a challenging problem, since even at low concentrations they can have drastic effects on the final habit. Industrially this causes problems with downstream processes such as filtration, processability and even storage. Conversely, structurally related additive molecules may be introduced to a system in order to mimic the effect of an impurity resulting in a beneficial effect on problematic crystal morphologies. The work presented here considers the design and use of tailor made additives on a nonhydrogen bonded crystal, benzophenone. This compound is typical of many agrochemical materials in that the major intermolecular interactions are of the nondirectional van der Waals type. Using crystal packing analysis a selection of additives has been chosen with the intent of specifically hindering certain directions of crystal growth. From an initial group of nine molecules two additives, 4ABP and 4MBP were found tobe particularly effective, both strongly hindering growth. Measured kinetic data suggests that these additives bind to steps in the growth spirals, drastically slowing growth of specific crystal faces altering the crystal morphology to a needle shape. Through nucleation experiments and product analysis the additives were shown to effect only crystal growth becoming incorporated into the crystal structure. Computational modelling of the binding of additives to the crystal surfaces of benzophenone has been used in an attempt to rationalise the experimental effects. In many cases calculated binding energies were in agreement with experimental observation. However, modified attachment energies did not match well with experimental observations.

Appartenant à la sous-famille des récepteurs couplés aux protéines G (RCPGs) apparentés àla rhodopsine et identifiés comme des cibles à fort potentiel thérapeutique, les récepteurs MT, et MT2à la mélatonine, 5-HT2C à la sérotonine et CB2 aux cannabinoïdes ont été étudiés par des approches insilico afin de mettre en évidence les déterminants structuraux critiques pour l'affinité, la sélectivité etl'activité pharmacologique de leurs ligands. Bénéficiant de données cristallographiques récentes,plusieurs états conformationnels de ces quatre récepteurs ont été modélisés en fonction du profilpharmacologique recherché. L'étude comparative de ces différents états conformationnels par dessimulations de dynamique moléculaire a permis de caractériser le rôle prépondérant joué par la boucleextracellulaire E2 et l'hélice 6 dans les mécanismes d'activation de ces RCPGs. Sur la base deméthodes chémoinformatiques, le criblage virtuel de ligands ciblant ces modèles tridimensionnels apermis de caractériser un modèle du récepteur 5-HT2C très spécifique de ligands agonistes inverses etd'identifier des touches pharmacologiques sur les récepteurs MTi et CB2. / Identified as highly relevant therapeutical targets, the MT, and MT2 melatonin receptors, the5-HT2C serotonin and the CB2 cannabinoid receptors, which belong to the rhodopsin-like G proteincoupledreceptors (GPCRs) subfamily, have been studied by in silico approaches in order to identifycritical structural features for the binding, the selectivity and the pharmacological activity of theirligands. Gaining by sottie recent crystallographic data, various conformational states of these fourreceptors have been modeled according to the expected pharmacological profile. The comparativestudy of these various conformational states by molecular dynamics simulations has led to emphasizethe crucial rôle of the E2 extracellular loop and hélix 6 in the activation mechanisms of these GPCRs.On the basis of chemoinformatic methods, the virtual ligand screening targeting these threedimensionalmodels has promoted the characterization of a 5-HT2C receptor model able to bindspecifically inverse agonist ligands and the identification of pharmacological hits targeting the MTiand CB2 receptors.

<div> <div> <div> <div> <div> <p>Polymer materials are receiving increased attention in the field of materials science, both in academia and industry, with its widespread application from commercial plastics to advanced biomaterials. These include composites in airplanes and automobiles, functional films on monitors in mobile phones and computers, as well as adhesive and coating materials in civil engineering. Despite significate efforts, the major questions and challenges in understanding key properties of polymer materials are still not solved. Such lack of understanding hinders advances in delicate design and controlling of polymers for advanced functional applications. The development of polymer science began with the pioneering work made by Flory and his coworkers at 1950s as commercial synthetic polymer industry started to develop and grow. During the following decades, experimental work guided by theoretical predictions had been the major contribution of our further understanding while the great challenges in experimental techniques at molecular level always blurred critical information in polymer materials. With enhanced ability in computational science, simulation starts to become an essential investigation method to provide thermodynamic insights at this molecular level. Along with great progress in properties prediction with improved accuracy, great challenges still exist in modeling processing of polymer systems, especially in accurate description of dynamic evolution incorporated with various processing conditions resulting macroscopic structural changes like carbon fiber processing from polyacrylonitrile (PAN) precursor in which crystalline regions represent more than 55% of the material by volume. In terms of crystallinity in polymers, with the heated debates over classical crystal-growth models, major questions and challenges are still not solved including the control and determination of molecular conformations and crystal structures as well as mesoscale morphologies, detailed understanding of melting and crystallization. It is clear that molecular scale investigations on crystal structure and crystallization mechanisms as well as predictive simulations of that will be a huge demand in the near future to explore mechanical, optical, and other physical properties in polymeric materials. </p> <p>The purpose of my dissertation is to summarize my major research contributions to our current understanding of crystalline polymers in the aspects of crystal structure determination and crystallization processes at molecular level, and to introduce our effort on simulation software development and indicate possible future directions in the field of molecular modeling of polymer crystals. Three major research topics will be included as the following </p> </div> </div> <div> <div> <p>1. Crystalline and pseudo-crystalline phases of polyacrylonitrile from molecular dynamics; 2. Novel mode of non-crystallographic branching in the initial stages of polymer fibril<br></p></div></div></div><div><div><div> <p>growth;<br> 3. Polymer crystal structure generator and analysis software (PolymerXtal). </p> </div> </div> </div> </div> </div>

Ce manuscrit présente une étude du procédé Supercritique Anti-Solvant (SAS) en combinant un travail expérimental et une étude de modélisation moléculaire. En comparaison aux méthodes traditionnelles de cristallisation en solution, le procédé SAS permet une baisse significative des quantités de solvants utilisées, un meilleur contrôle des caractéristiques des poudres générées ainsi qu’une plus grande sélectivité polymorphique. De nombreuses études expérimentales ou de modélisation numérique ont permis une meilleure compréhension de ce procédé, mais certains aspects, liés aux mécanismes de cristallisation sous pression, demeurent moins discutés. Par une investigation de ces mécanismes, l’objectif de ce travail a été de développer et de valider des méthodes permettant un meilleur contrôle du faciès des poudres générées et de la forme du polymorphe. De ces caractéristiques dépendent certaines propriétés des cristaux, telles que leur cinétique de dissolution ou encore leur stabilité physique et chimique, particulièrement importante pour le domaine pharmaceutique / This manuscript presents a study of the Supercritical Anti-Solvent (SAS) process by combining an experimental work and a molecular modeling study. Compared with conventional crystallization methods in liquid solutions, the SAS process allows a significant decrease in the amounts of solvents used, a better control of generated powder characteristics as well as a greater polymorphic selectivity. Many experimental studies or numerical modeling have allowed a better understanding of this process, but some aspects, related to crystallization mechanisms under pressure, remain less discussed. Through an investigation of these mechanisms, the objective of this work was to develop and validate methods allowing a better control of the crystal habit and of the polymorphic form of generated powders. Certain crystal properties depend on these characteristics, such as their dissolution kinetics or their physical and chemical stability, which are particularly important in the pharmaceutical field