La dispersion dans des milieux poreux homogènes (empilements de grains) a été étudiée par des mesures par résonance magnétique nucléaire (RMN) et des simulations de marches aléatoires dans un réseau de pores. La RMN permet de mesurer l’ensemble des déplacements des molécules d’eau durant un temps tΔ, et d’obtenir propagateurs et moments caractéristiques. L’évolution temporelle du second moment σ (σ2 ∝ taΔ) permet de caractériser de manière précise le régime de dispersion des molécules (Gaussien ou anormal). Des mesures pour des écoulements de 15 < Pe < 45 dans un empilement de grains de 30μm ont permis d’observer une dispersion anormale faiblement super-dispersive (a = 1.17) en écoulement saturé et une augmentation progressive du caractère super-dispersif avec la diminution de la saturation en eau (jusqu’à a = 1.5 pour 42 %)lors d’une co-injection stationnaire eau-huile. En écoulement saturé, les propagateurs et courbes de percée sont quasi-gaussiennes, tandis qu’en écoulement insaturé, les propagateurs sont asymétriques et les courbes de percée présentent des trainées aux grands temps. Dans ces conditions, on montre que la dispersion anormale observée est mieux décrite par des lois stables de Lévy que par des lois gaussiennes. Des simulations de marche aléatoire ont été réalisées dans un réseau de pores extrait d’un milieu poreux réel par imagerie microscanner.Elles permettent d’obtenir les mêmes informations que la RMN, les marcheurs se déplaçant par advection et diffusion. Ces simulations montrent l’existence d’une stagnation non observée dans les expériences, montrant que la simplification du réseau poreux est trop importante et empêche de reproduire certains aspects du champ de vitesses détecté par la RMN. Toutefois, l’évolution temporelle du second moment a également un caractère super-dispersif à temps long à 100 % de saturation / We investigated dispersion in homogeneous porous media (grain packs) by nuclear magnetic resonance (NMR) measurements and random walk simulations in pore networks. We measured water molecules displacements during a time interval tΔ by NMR measurements, which allows us to obtain propagators and charateristic cumulants of displacements such as the mean square displacement σ. The evolution of the cumulant σ as a function of time tΔ (σ2 ∝ taΔ) is a very sensitive test of Gaussian behaviour compared to the analysis of the shape of propagators. In a homogeneous 30μm grain pack and low Peclet numbers (15 < Pe < 45), we observed weak super dispersion in saturated conditions (a = 1.17) and gradually stronger super-dispersionas the water saturation decreases (up to a = 1.5 for 42 %) during steady-state oil-water two phase flow. Insaturated conditions, propagators and breakthrough curves are Gaussian or nearly Gaussian, whereas in two phase conditions, propagators are non symmetric and breakthrough curves show thick tails at long time. Weshow that the anomalous dispersion observed is better explained by Lévy stable laws (asymetric for longitudina ldispersion, and symetric for transverse dispersion) than by Gaussian laws. Random walk simulations were performed in a pore network constructed using high resolution images of the grain pack. They allow us to obtain the same informations than the NMR, with walkers submitted to diffusive and advective effects. The simulations show the existence of an anomalous stagnation not observed in experiments, highlighting the oversimplification of the pore network that prevent reproducing some aspects of the velocity field detected by NMR. However, the simulations indicate similarly a super-dispersion at long time in saturated conditions

Com a crescente disponibilidade de luz síncrotron tem-se utilizado cada vez mais a dispersão anômala (f´) no estudo de materiais. Apesar dos vários métodos já empregados na medida do fator de espalhamento atômico, ainda é pequeno o número de elementos abrangidos. Este trabalho apresenta uma contribuição à ampliação da gama de elementos mensuráveis através de um método interferométrico diferencial para a determinação experimental de f´. A sua demonstração é feita através de medidas em Selênio obtendo-se o mesmo nível de precisão do método &#933-&#933/2 usado em trabalhos anteriores. Esses dois métodos são criticados quanto ao limite de erro atingível devido à sua natureza não-absoluta. Uma técnica de preparação de amostras para materiais de baixo ponto de fusão é explorada, permitindo determinar f´absolutamente, com erro de 0,03 elétron. A medida do fator de espalhamento atômico através do desvio angular entre as reflexões de Bragg e Laue é proposta e testada com resultados de qualidade bastante inferior às técnicas interferométricas. A construção de interferômetros de raios X estáveis e de alto contraste é estudada quanto à seleção dos monocristais de silício e tratamento após o corte do dispositivo, atingindo-se contraste de 80% com estabilidade de uma milifranja por hora. / The use of anomalous dispersion (f\') in the study of materials has been increasing with the availability of synchrotron radiation. Although many methods have been designed for the measurement of the atomic scattering factor, the number of elements the cover is still small. This work presents a contribution to the widening of the range of measurable elements through a Differential interferometric method for the determination of f´. Its demonstration is achieved by measurements on Selenium, reaching the same level of accuracy obtained by the &#933-&#933/2 method used in previous work. These two methods are criticized in terms of the achievable errors due to its non-absolute nature. A technique for the preparation of samples with low melting point materials is developed, allowing the determination of f´ absolutely, with an error of 0,03 electron. The measurement of atomic scattering factor through the angular offset between Bragg and Laue reflections is proposed and tested, with low quality results compared to interferometric methods. The construction of high stability and good contrast X-ray interferometers is studied in terms of silicon single crystal selection and its treatment after device cutting. 80% contrast was achieved with a stability of one milifringe per hour.

The crystal structures of two thermostable carboxylesterase Est30 and Est55 from Geobacillus stearothermophilus were determined to help understand their functions and applications in industry or medicine. The crystal structure of Est30 was determined at 1.63 Å resolution by the multiple anomalous dispersion method. The two-domain Est30 structure showed a large domain with a modified alpha/beta hydrolase core including a seven, rather than an eight-stranded beta sheet, and a smaller cap domain comprising three alpha helices. A 100 Da tetrahedral ligand, propyl acetate, was observed to be covalently bound to the side chain of Ser94 in the catalytic triad. This ligand complex represents the first tetrahedral intermediate in the reaction mechanism. Therefore, this Est30 crystal structure will help understand the mode of action of all enzymes in the serine hydrolase superfamily. Est55 is a bacterial homologue of the mammalian carboxylesterases involved in hydrolysis and detoxification of numerous peptides and drugs and in prodrug activation. Est55 crystals were grown at pH 6.2 and pH 6.8 and the structures were determined at resolutions of 2.0 and 1.58 Å respectively. Est55 folds into three domains, a catalytic domain, an α/β domain and a regulatory domain. This structure is in an inactive form; the side chain of His409, one of the catalytic triad residues, is pointing away from the active site. Moreover, the adjacent Cys408 is triply oxidized and lies in the oxyanion hole, which would block the entry of substrate to its binding site. This structure suggested a self-inactivation mechanism, however, Cys408 is not essential for enzyme activity. Mutation of Cys408 showed that hydrophobic side chains at this position were favorable, while polar serine was unfavorable for enzyme activity. Both Est30 and Est55 were shown to hydrolyze the prodrug CPT-11 into the active form SN-38. Therefore, Est30 and Est55 are potential candidates for use with irinotecan in cancer therapy. The catalytic efficiency (kcat/Km) of Est30 is about 10-fold lower than that of Est55. The effects of the Cys408 substitutions on Est55 activity differed for the two substrates, p-NP butyrate and CPT-11. Mutant C408V may provide a more stable form of Est55.

Com a crescente disponibilidade de luz síncrotron tem-se utilizado cada vez mais a dispersão anômala (f´) no estudo de materiais. Apesar dos vários métodos já empregados na medida do fator de espalhamento atômico, ainda é pequeno o número de elementos abrangidos. Este trabalho apresenta uma contribuição à ampliação da gama de elementos mensuráveis através de um método interferométrico diferencial para a determinação experimental de f´. A sua demonstração é feita através de medidas em Selênio obtendo-se o mesmo nível de precisão do método &#933-&#933/2 usado em trabalhos anteriores. Esses dois métodos são criticados quanto ao limite de erro atingível devido à sua natureza não-absoluta. Uma técnica de preparação de amostras para materiais de baixo ponto de fusão é explorada, permitindo determinar f´absolutamente, com erro de 0,03 elétron. A medida do fator de espalhamento atômico através do desvio angular entre as reflexões de Bragg e Laue é proposta e testada com resultados de qualidade bastante inferior às técnicas interferométricas. A construção de interferômetros de raios X estáveis e de alto contraste é estudada quanto à seleção dos monocristais de silício e tratamento após o corte do dispositivo, atingindo-se contraste de 80% com estabilidade de uma milifranja por hora. / The use of anomalous dispersion (f\') in the study of materials has been increasing with the availability of synchrotron radiation. Although many methods have been designed for the measurement of the atomic scattering factor, the number of elements the cover is still small. This work presents a contribution to the widening of the range of measurable elements through a Differential interferometric method for the determination of f´. Its demonstration is achieved by measurements on Selenium, reaching the same level of accuracy obtained by the &#933-&#933/2 method used in previous work. These two methods are criticized in terms of the achievable errors due to its non-absolute nature. A technique for the preparation of samples with low melting point materials is developed, allowing the determination of f´ absolutely, with an error of 0,03 electron. The measurement of atomic scattering factor through the angular offset between Bragg and Laue reflections is proposed and tested, with low quality results compared to interferometric methods. The construction of high stability and good contrast X-ray interferometers is studied in terms of silicon single crystal selection and its treatment after device cutting. 80% contrast was achieved with a stability of one milifringe per hour.

Second harmonic (SH) microscopy represents a powerful tool for the investigation of crystalline systems, such as ferroelectrics and their domain walls (DWs). Under the condition of normal dispersion, i.e., the refractive index at the SH wavelength is larger as compared to the refractive index at the fundamental wavelength, n(2ω) . n(ω), bulk crystals will generate no SH signal. Should the bulk, however, contain DWs, an appreciable SH signal will still be detectable at the location of DWs stemming from the Čerenkov mechanism. In this work, we demonstrate both how SH signals are generated in bulk media and how the Čerenkov mechanism can be inhibited by using anomalous dispersion, i.e., n(ω) . n(2ω). This allows us to quantitatively estimate the relative strength of the Čerenkov compared to other SH contrast mechanisms in DWs, such as the interference contrast. The results are in agreement with previous experiments based on the geometric separation of the signals. Due to the observed, strong Čerenkov contrast, such signal contributions may not be neglected in polarimetry studies of ferroelectric DWs in the future.

Ce travail a pour but de proposer des outils visant `a comparer des résultats exp´erimentaux avec des modèles pour la dispersion de traceur en milieu poreux, dans le cadre de la dispersion anormale.Le “Mobile Immobile Model” (MIM) a été à l’origine d’importants progrès dans la description du transport en milieu poreux, surtout dans les milieux naturels. Ce modèle généralise l’quation d’advection-dispersion (ADE) e nsupposant que les particules de fluide, comme de solut´e, peuvent ˆetre immo-bilis´ees (en relation avec la matrice solide) puis relˆachées, le piégeage et le relargage suivant de plus une cin´etique d’ordre un. Récemment, une version stochastique de ce modèle a ´eté proposée. Malgré de nombreux succès pendant plus de trois décades, le MIM reste incapable de repr´esenter l’´evolutionde la concentration d’un traceur dans certains milieux poreux insaturés. Eneffet, on observe souvent que la concentration peut d´ecroˆıtre comme unepuissance du temps, en particulier aux grands temps. Ceci est incompatible avec la version originale du MIM. En supposant une cinétique de piégeage-relargage diff´erente, certains auteurs ont propos´e une version fractionnaire,le “fractal MIM” (fMIM). C’est une classe d’´equations aux d´eriv´ees par-tielles (e.d.p.) qui ont la particularit´e de contenir un op´erateur int´egral li´e`a la variable temps. Les solutions de cette classe d’e.d.p. se comportentasymptotiquement comme des puissances du temps, comme d’ailleurs cellesde l’´equation de Fokker-Planck fractionnaire (FFPE). Notre travail fait partie d’un projet incluant des exp´eriences de tra¸cageet de vélocimétrie par R´esistance Magn´etique Nucl´eaire (RMN) en milieuporeux insatur´e. Comme le MIM, le fMIM fait partie des mod`eles ser-vant `a interpréter de telles exp´eriences. Sa version “e.d.p.” est adapt´eeaux grandeurs mesur´ees lors d’exp´eriences de tra¸cage, mais est peu utile pour la vélocimétrie RMN. En effet, cette technique mesure la statistiquedes d´eplacements des mol´ecules excit´ees, entre deux instants fixés. Plus précisément, elle mesure la fonction caractéristique (transform´ee de Fourier) de ces d´eplacements. Notre travail propose un outil d’analyse pour ces expériences: il s’agit d’une expression exacte de la fonction caract´eristiquedes d´eplacements de la version stochastique du mod`ele fMIM, sans oublier les MIM et FFPE. Ces processus sont obtenus `a partir du mouvement Brown-ien (plus un terme convectif) par des changement de temps aléatoires. Ondit aussi que ces processus sont des mouvement Browniens, subordonnéspar des changements de temps qui sont eux-mˆeme les inverses de processusde L´evy non d´ecroissants (les subordinateurs). Les subordinateurs associés aux modèles fMIM et FFPE sont des processus stables, les subordinateursassoci´es au MIM sont des processus de Poisson composites. Des résultatsexp´erimenatux tr`es r´ecents on sugg´er´e d’´elargir ceci `a des vols de L´evy (plusg´en´eraux que le mouvement Brownien) subordonnés aussi.Le lien entre les e.d.p. fractionnaires et les mod`eles stochastiques pourla sous-diffusion a fait l’objet de nombreux travaux. Nous contribuons `ad´etailler ce lien en faisant apparaˆıtre les flux de solut´e, en insistant sur une situation peu ´etudiée: nous examinons le cas o`u la cinétique de piégeage-relargage n’est pas la mˆeme dans tout le milieu. En supposant deux cinétiques diff´erentes dans deux sous-domaines, nous obtenons une version du fMIMavec un opérateur intégro-diff´erentiel li´e au temps, mais dépendant de la position.Ces r´esultats sont obtenus au moyen de raisonnements, et sont illustrés par des simulations utilisant la discrétisation d’intégrales fractionnaires etd’e.d.p. ainsi que la méthode de Monte Carlo. Ces simulations sont en quelque sorte des preuves numériques. Les outils sur lesquels elles s’appuient sont présentés aussi. / We propose tools for to compare experimental data and models for anomalousdispersion in porous media.The “Mobile Immobile Model” (MIM) significantly improved the descrip-tion of mass transport in natural porous media. This model generalizes theadvection-dispersion equation (ADE) by assuming that fluid and solute parti-cles can be found in mobile on immobile states, exchanging matter accordingto first order kinetics. Moreover, it has a stochastic version. Nevertheless,the original MIM does not represent the power-law decrease of some break-through curves observed in some media, better described by a fractionalversion, the “fractal MIM” (fMIM) which assumes a different kinetics. Theacronym “fMIM” denotes partial differential equations (p.d.e.) involving afractional integral with respect to time, having solutions falling-off as powerof times, asymptotically. It keeps in similarity with the fractional Fokker-Planck equation (FFPE). As this equation, the fMIM describes the evolutionof the probability density function of stochastic processes, namely Brownianmotion sujected to a time change that is the hitting time of a stable sub-ordinator, strictly stable or not, according FFPE or fMIM is considered.Using probabilistic arguments and numerical simulation, we extend this re-sult to the case when the transport parameters and the time scales of thetime change vary in space. P.d.es are well suited for comparing with tracer tests data. Yet, they arenot very useful to discuss signals recorded by pulsed field gradient (PFG)nuclear magnetic resonance (NMR), a technique which measures the char-acteristic function (Fourier transform) of molecular displacements betweentwo fixed instants. For to process such data, we derive an expression of thecharacteristic function of the displacements of Brownian motions subordi-nated by the hitting times of stable subordinators, i.e. of processes whosedensity satisfies FFPE of fMIM. We also consider time changes that are hit-ting times of composite Poisson processes (CPP), which correspond to theoriginal version of the MIM.

No description available.

No description available.

N-glycosylation is the most common eukaryotic post-translational modification, impacting on protein stability, folding, and protein-protein interactions. More broadly, N-glycans play biological roles in reaction kinetics modulation, intracellular protein trafficking, and cell-cell communications. The machinery responsible for the initial stages of N-glycan assembly and processing is found on the membrane of the endoplasmic reticulum. Following N-glycan transfer to a nascent glycoprotein, the enzyme Processing α-Glucosidase I (GluI) catalyzes the selective removal of the terminal glucose residue. GluI is a highly substrate-specific enzyme, requiring a minimum glucotriose for catalysis; this glycan is uniquely found in biology in this pathway. The structural basis of the high substrate selectivity and the details of the mechanism of hydrolysis of this reaction have not been characterized. Understanding the structural foundation of this unique relationship forms the major aim of this work. To approach this goal, the S. cerevisiae homolog soluble protein, Cwht1p, was investigated. Cwht1p was expressed and purified in the methyltrophic yeast P. pastoris, improving protein yield to be sufficient for crystallization screens. From Cwht1p crystals, the structure was solved using mercury SAD phasing at a resolution of 2 Å, and two catalytic residues were proposed based upon structural similarity with characterized enzymes. Subsequently, computational methods using a glucotriose ligand were applied to predict the mode of substrate binding. From these results, a proposed model of substrate binding has been formulated, which may be conserved in eukaryotic GluI homologs.

N-glycosylation is the most common eukaryotic post-translational modification, impacting on protein stability, folding, and protein-protein interactions. More broadly, N-glycans play biological roles in reaction kinetics modulation, intracellular protein trafficking, and cell-cell communications. The machinery responsible for the initial stages of N-glycan assembly and processing is found on the membrane of the endoplasmic reticulum. Following N-glycan transfer to a nascent glycoprotein, the enzyme Processing α-Glucosidase I (GluI) catalyzes the selective removal of the terminal glucose residue. GluI is a highly substrate-specific enzyme, requiring a minimum glucotriose for catalysis; this glycan is uniquely found in biology in this pathway. The structural basis of the high substrate selectivity and the details of the mechanism of hydrolysis of this reaction have not been characterized. Understanding the structural foundation of this unique relationship forms the major aim of this work. To approach this goal, the S. cerevisiae homolog soluble protein, Cwht1p, was investigated. Cwht1p was expressed and purified in the methyltrophic yeast P. pastoris, improving protein yield to be sufficient for crystallization screens. From Cwht1p crystals, the structure was solved using mercury SAD phasing at a resolution of 2 Å, and two catalytic residues were proposed based upon structural similarity with characterized enzymes. Subsequently, computational methods using a glucotriose ligand were applied to predict the mode of substrate binding. From these results, a proposed model of substrate binding has been formulated, which may be conserved in eukaryotic GluI homologs.