The poultry-to-human transmission of the influenza virus and the recent H1Nl influenza pandemic have become major concerns worldwide. The nucleoprotein (NP) of influenza virus binds the RNA genome and plays essential role in transcription and replication during the virus life cycle. / The study leads to a better understanding towards the RNP organization of influenza virus and provides information for the future design of anti-influenza agents. / We have also shown, by RNP reconstitution assay and co-immunoprecipitation, that the interaction between NP and PB2 is crucial for the proper functioning of the RNP. The functional association of NP and PB2 requires either the PB2 host-determining residue lysine-627 or arginine-630 with the latter involving NP arginine-150 also. Using SPR, we have demonstrated that both residues take part in the direct protein-protein interaction, without the involvement of RNA. These results suggest a dual interaction mechanism between NP and PB2. This may confer replication advantages to the virus, as either one can give an active RNP and explains the increased virulence of avian influenza viruses carrying the E627K mutation in mammalian cells. In addition, our findings identify the NP-PB2 interacting surface, with the PB2 627/630 region facing the RNA binding groove of NP. / We have determined the 3.3 A crystal structure of H5N1 NP, which is composed of head and body domains and a tail loop. Using surface plasmon resonance (SPR), we found the basic loop (residues 73-91) and arginine-rich groove, but mostly a protruding element centering at R174 and R175, to be important in RNA binding. Ribonucleoprotein (RNP) reconstitution assay with these multiple-point and deletion mutants indicate their functional importance towards the transcription-replication activities of the virus polymerase. Single-point mutations at these concerned regions do not have a significant effect on their RNP activities, suggesting that NP mediates RNA-binding through multiple residues. / Ng, Ka Leung. / Adviser: Pang Chui Shaw. / Source: Dissertation Abstracts International, Volume: 73-06, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 121-136). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

This qualitative study explores midwifery practice during the second stage of labour focusing specifically on whether midwives adopt a directed or physiological approach to maternal pushing. It was undertaken against the backdrop of research findings suggesting that there is no proven benefit to directing a woman's pushing efforts but anecdotal evidence suggests that this remains a routine and accepted part of midwifery practice in the United Kingdom (UK). Semi- structured interviews were undertaken with ten midwives who had recent experience of caring for women during the second stage of labour, ten women who had recently given birth and four obstetricians. A form of thematic analysis was undertaken. Findings were viewed through a lens of critical social theory (CST) and drew on feminist principles to provide a deeper understanding of the emergent themes. Findings indicated that a directed approach to second stage pushing was the norm in this UK Maternity Unit and was deeply embedded within the cultural context of what it meant to be a midwife that involved ' doing' rather than 'being'. Reasons explaining why midwives continue to use directed pushing were grouped into themes; ' time passing and watching the clock' 'different worlds' , 'different women', 'midwives take charge', 'growth of confidence and changing practice' and 'conflict'. When viewed from a CST perspective midwives undertaking directed pushing is seen as an example of institutionalised oppressive behaviour symbolising the way in which knowledge and rationality are disregarded in favour of a risk averse practice that is paradoxically the opposite of what evidence recommends. Midwives are identified as being oppressed by the dominant biomedical model to the extent that they do not view directed pushing as an intervention. In order to promote a more physiological approach with its' associated benefits, a return to a social model of midwifery with a focus on salutogenesis rather than pathogenesis is called for. Recommendations for midwifery education, practice and research are provided in order to support the transformational shift in midwifery culture that is needed if such a change is to become a reality.

Coenzima Q (CoQ) é uma molécula de função essencial na transferência de elétrons da cadeia respiratória mitocondrial. Em Saccharomyces cerevisiae , a CoQ é constituída por um anel de benzeno associado a uma cadeia poliprenil, com 6 unidades de repetição, sendo por isso também denominada CoQ6 ou Q6. Ao todo já foram identificados treze genes (COQ1 COQ11, ARH1 e YAH1) nucleares necessários para biossíntese da CoQ. A maioria dos produtos Coq estão fisicamente associados em um complexo biossintético ancorado na membrana mitocondrial interna. Neste projeto, tentamos descrever resíduos relevantes de Coq3p e Coq7p aliando análises de bioinformática com testes fenotípicos para balizamento funcional. Coq7p é uma proteína com dois centros de ferro com íons carboxilato e catalisa a hidroxilação de demetoxi-Q6 (DMQ6). Neste estudo, indicamos um grupo de resíduos que modulam a atividade e a estabilidade de Coq7p: D53, R57, V111 e S114. Enquanto R57, V111 e S114 são resíduos muito conservados, V111 e S114 estão correlacionados em comunidades de coevolução. Aqui, demonstramos também que o duplo mutante S114A, V111G e o mutante S114E apresentam deficiência respiratória em temperatura não permissiva, além de acumularem o intermediário DMQ6 e sintetizarem baixas quantidades de Q6, concluindo assim que o fosmimético S114E inibe a atividade Coq7p. Dessa forma, propomos que a fosforilação do resíduo S114 promove o deslocamento de uma alça entre as hélices 2 e 3, afetando assim a atividade do centro catalítico Coq7p. Por sua vez, Coq3p atua como uma metiltransferase, catalisando diferentes passos durante a biossíntese da CoQ. Aqui, identificamos resíduos que colaboram para a atividade funcional de Coq3p: E123, S125, C131, G133, G134, H165, D203, E219, K258 e S262. Mutantes carregando as alterações E123A, H165A, D203A, E219A, K258A e S62A apresentam discreto crescimento respiratório e expressão de Coq3p similares à da linhagem selvagem, além de acumularem baixas quantidades de Q6. Enquanto C131, G133 e G134 são resíduos altamente conservados, localizados em uma alça no espaço entre fitas beta, no provável sítio ativo da proteína, mutantes C131A, G133A e G134A se superexpressos apresentam crescimento respiratório em meio contendo fonte de carbono não fermentável, além de acumularem Q6 compatíveis com os níveis de expressão proteica. Propomos assim um modelo para Coq3p, tendo os resíduos C131, G133 e G134 como centro catalítico de Coq3p. / Coenzyme Q (CoQ) is a molecule of essential function in the transfer of electrons of the mitochondrial respiratory chain. In saccharomyces cerevisiae , CoQ is constituted by a benzene ring associated with a polyprenyl chain with 6 repetition units, being therefore also denominated CoQ6 or Q6. Thirteen nuclear genes have already been identified (COQ1 COQ11, ARH1 and YAH1) required for coenzyme Q biosynthesis. Most of Coq products are physically associated in a biosynthetic complex anchored at the mitochondrial internal membrane. In this project, we identified Coq3p and Coq7p residues relevant for their respective role in CoQ synthesis combining bioinformatics analyzes with phenotypic tests for functional mapping. Coq7p is a carboxylate-bridged di-iron protein that catalyzes the hydroxylation of demetoxy-Q6 (DMQ6), the last monooxygenase step in the synthesis of CoQ. In this study, we found a group of residues that modulate the activity and stability of Coq7p: D53, R57, V111 and S114. While R57, V111 and S114 are highly conserved residues, V111 and S114 are correlated in communities of coevolution. We also demonstrate that the double mutant S114A, V111G and the mutant S114E have respiratory deficiency at non-permissive temperature, in addition to accumulating of the intermediate DMQ6 and low amounts of Q6, thus concluding that phosmimetic S114E inhibits the activity of Coq7p. Hence, we propose that the phosphorylation of S114 is required to move a loop between helices 2 and 3, thus affecting the activity of the catalytic center Coq7p. For its part, Coq3p acts as a methyltransferase, catalyzing different steps during biosynthesis of CoQ. Here we identified residues that collaborate for functional activity of Coq3p: E123, S125, C131, G133, G134, H165, D203, E219, K258 and S262. Mutants E123A, H165A, D203A, E219A, K258A and S62A, have mild respiratory growth, and expression of Coq3p levels similar to the wild strain, in addition to accumulating low amounts of Q6. While C131, G133, and G134 are residues highly conserved, located in a loop in the space between beta sheets, the overexpression of the mutants C131A, G133A and G134A present respiratory growth in medium containing non-fermentable carbon source, in addition to accumulate Q6 compatible with the levels of protein expression. We propose a model for Coq3p, with residues C131, G133 and G134 as part of Coq3p catalytic center.

Neste trabalho foram estudadas algumas formas de relação entre séries temporais multivariadas. Discutiu-se, inicialmente, a função de coerência, uma função análoga a função de correlação(que é dada no domínio do tempo) calculada no domínio da freqüência. Foram estudadas também as funções de coerência parcial e coerência parcial direcionada. A função de coerência parcial mede a relação entre duas componentes de uma série multivariada, isolados os efeitos de outra série. Em linhas gerais, a Coerência Parcial Direcionada pode ser interpredata como a decomposição da coerência parcial a partir de modelos autoregressivos multivariados. Esse conceito pode ser interpretado como uma representação do conceito de causalidade de Granger no domínio da freqüência. Finalmente, foram aplicadas as funções acima em dois conjuntos de dados: um modelo VAR(1) trivariado simulado e dados de medições de eletroencefalograma. / In this work we studied relationships between multivariate time series. We discussed the coherence function, a function similar to the correlation function(calculated in time domain) in frequency domain. Next, we discussed partial coherence and partial directed coherence. The partial coherence measures the relationship between two components of a multivariate time series, after removing the influence of another time series. Generally, the partial directed coherence can be interpreted as the decompositioin of the partial coherence from multivariate autoregressive models. We can interpret this function as a representation of the Granger causality concept in frequency domain. Finally, we applied these concepts in two situations: a simulated VAR(1) model and an electroencefalogram database.

Cette thèse se concentre sur la problématique suivante : comment assister un utilisateur novice pour l'aider à obtenir un dessin de son graphe qui soit adapté à ses besoins ? En effet, les méthodes de dessins actuelles, très nombreuses, nécessitent une grande expertise pour obtenir un dessin de bonne qualité. Or, par manque d'expertise, les utilisateurs novices ne peuvent pour l'instant pas produire des dessins d'une telle qualité à partir de leurs données. La solution proposée consiste à mettre en place un système interactif proposant à l'utilisateur différents dessins pour un même graphe afin qu'il obtienne un résultat qui réponde correctement à ses besoins. Ce système se base sur un algorithme de force modifié utilisé par un système d'algorithme génétique hautement modulable. L'objectif de la modification apportée à l'algorithme de dessin étant de pouvoir générer plusieurs dessins intéressants pour un même graphe. / The main objective of this thesis is to deal with assisting a novice user in drawinga graph which conforms to his/her needs. Currently, a lot of different methods for graph drawing exist, but they need an high level of expertise to be efficiently used. However, novice users don't have this kind of expertise, and thus they usually use the most common drawing methods. We design a solution to deal with this problem using an interactive system which generate several different drawings for a graph and then let the user choose which best conform to his/her constraints. This system is based on a modified force-directed algorithm controlled by a highly parameterisable genetic algorithm. The aim of the modification applied to the force-directed algorithm is to generate several different and interesting drawings of the same graph, by setting the parameters for each vertex (instead of global graph values).

L'application des techniques de séquençage à haut débit dans les dernières années a conduit à l'obtention de la séquence de génomes complets de plusieurs organismes. Le développement de nouveaux outils de génétique inverse était donc souhaitable afin de faire un usage optimal des données accumulées. Les nucléases hautement spécifiques représentent un outil unique pour induire des modifications ciblées du génome in vivo. L'induction d'une cassure double brin dans l'ADN est réparée soit par la voie de jonction d’extrémités nonhomologues soit par la voie très fidèle de la recombinaison homologue. Le ciblage d'un locus précis avec une nucléase spécifique stimule fortement la réparation de l'ADN, qui peut être utilisé pour induire des modifications ciblées dans le génome. Dans ma thèse, je vise à fournir une preuve de prinicipe pour l'utilisation des méganucléases et des transcription activator like effector nucléases (TALENs), deux classes communes de nucléases très spécifiques, comme nouveaux outils d'édition du génome chez le medaka, Oryzias latipes, et le poisson zèbre, Danio rerio. J'ai d’abord trouvé les conditions optimales d'utilisation de ces nucléases dans nos modèles de poissons. J'ai à cette occasion également développé une méthode très sensible et rapide pour la détection de modifications génomiques ciblées. J'ai ensuite induit des mutations au sein de trois gènes différents chez le poisson zèbre avec des TALENs. Les mutations dans le gène col6a1 ont conduit à la mise en évidence pour la première fois d’une technique de modification d’un site d’épissage d’un gène de poisson zèbre à l’aide d’une nucléase ciblée. Ce travail nous a permis d’établir une lignée de poisson avec une mutation dans le collagène VI alpha 1 qui est similaire à une mutation fréquemment trouvée chez les patients humains atteints de myopathie de Bethlem. De même, j’ai pu induire des mutations dans le gène nle1 du poisson zèbre qui vont permettre la mise en place de lignées de poissons mutants de ce gène. En outre, j'ai pu montrer qu’un nouveau type de nucléase, une TALEN Compact, était actif sur une cible chromosomique chez le poisson zèbre. En conclusion, les études que j'ai effectuées ont apporté la preuve de principe pour l'activité de TALENs et Compact TALENs ainsi que la première démonstration de modification de l'épissage à l’aide d’une TALEN chez le poisson zèbre et ont abouti à la mise en place d'une lignée de poisson dont le phénotype est proche d’un syndrome humain ouvrant la voie à la création de modèles pour d’autres mutations de ce type. Pour finir, je discute dans cette thèse des conditions permettant un usage le plus efficace des nucléases ciblées pour la génération de mutants et l’édition du génome. / The application of high throughput sequencing techniques in the recent years has led to obtaining the full genome sequences of many organisms. The development of novel tools for reverse genetics was thus desirable to make optimal use of the accumulated data. Site directed nucleases represent a unique platform to induce targeted genome modifications in vivo. Targeting a precise locus with a highly specific nuclease stimulates DNA repair, which can be harnessed for genome editing. Induction of a double strand break in DNA is repaired by either the error prone pathway of nonhomologous end joining or the high fidelity pathway of homologous recombination in the cell. Both mechanisms can be used to insert foreign DNA into the genome of the host. In my thesis, I aimed to provide proof of principle for the use of meganucleases and transcription activator like effector nucleases (TALENs), two common classes of site directed nucleases, as novel tools for genome editing in medaka, Oryzias latipes, and zebrafish, Danio rerio. During the first years of my thesis, I found the optimal conditions to use these nucleases in our fish models. I also developed a very sensitive and rapid method for detection of targeted genome modifications. I then induced mutations at three different endogenous loci in zebrafish with TALENs. The mutations in the col6a1 gene led to the first demonstration of splicing site modification in zebrafish using a TALE nuclease. This allowed the establishment of a fish line with a mutation in type VI collagen alpha 1 chain homologous to one mutation frequently found in human patients with Bethlem myopathy. Then I generated mutations in the nle1 gene which are heritable and from which establishment of mutant fish lines is in progress. In addition, by using the method for detection of targeted genome modifications I developed, I showed that a novel type of nuclease, a Compact TALEN, was active on a chromosomal target in zebrafish. In conclusion, the studies I performed provided proof of principle for the activity of TALENs and Compact TALENs as well as the first demonstration of TALEN-Mediated modification of splicing in zebrafish and resulted in the establishment of a fish line with mutated collagen VI. Induction of heritable mutations in the nle1 gene in zebrafish was also confirmed. Additionally, I proved that the choice of expression vector is crucial for the synthesis of active site directed nucleases for use in fish and established a novel efficient method for detection of targeted genomic mutations.

Esta dissertação apresenta o desenvolvimento, a validação e a aplicação de algoritmos para inferência de conectividade neural em registros de EEG contendo potenciais evento-relacionados (ERP). Os sinais foram caracterizados via modelos auto-regressivos multivariados (MVAR) e empregou-se a coerência parcial direcionada (PDC) no estudo das relações de causalidade entre eles. Certas características dos ERPs, como sua transitoriedade intrínseca e as múltiplas repetições em experimentos, levaram ao desenvolvimento de novos algoritmos, como a estimação de modelos conjuntos a partir de vários segmentos de sinal e um procedimento em janela deslizante capaz de descrever a evolução temporal da estatística dos sinais de interesse. Ademais, mostrou-se a possibilidade de estender os resultados da análise assintótica da estatística da PDC ao caso multi-trecho, tornando possível o estudo de sua significância estatística sem recorrer a procedimentos de reamostragem. Os algoritmos foram validados em exemplos com neural mass models, modelos não-lineares capazes de gerar sinais com características muito semelhantes a sinais de EEG reais, e aplicados a uma base de dados pública contendo resultados de experimentos com ratos. / This dissertation presents the development, validation, and application of algorithms for inferring neural connectivity in EEG signals containing event-related potentials (ERP). The time series were described via multivariate auto-regressive models (MVAR) and partial directed coherence (PDC) was used to study causal relations between them. Certain features of the ERPs, such as their transitory behavior and the existence of multiple trials in an experiment, lead to the development of a new algorithm capable of estimating a joint model from multiple segments and a sliding-window procedure for describing the nonstationarity behavior of the signals of interest. Furthermore, the possibility of extending the asymptotic results for PDC\'s statistics to the multi-trial case was demonstrated, allowing, therefore, the study of its statistical significance without recurring to resampling methods. The algorithms were validated in examples with neural mass models, non-linear models capable of generating signals with features very similar to real EEG recordings, and then applied to a publicly available dataset of experiments in rats.

Os programas de computador têm apoiado o estudo de sistemas biomédicos em que um volume considerável de dados são empregados. Na biomecânica, a análise das influências entre as articulações pode melhorar o conhecimento das lesões relacionadas à corrida associadas ao uso excessivo durante a atividade de corrida. Compreender os padrões de interação entre diferentes articulações anatômicas, durante o movimento, pode contribuir para o aprimoramento de programas de treinamento, reabilitação e prevenção a lesões. Neste trabalho, um software personalizado foi desenvolvido para implementar a Coerência Parcial Direcionada (PDC), uma abordagem no domínio da freqüência da Causalidade de Granger (GC), adequado às especificidades da fisioterapia. Com entradas independentes e padronizadas, modularização e parametrização, as rotinas investigaram a direção de interação entre diferentes canais, registrando e salvando arquivos intermediários. Separados nos três planos anatômicos, sagital, frontal e transverso, foram utilizados dados cinemáticos para analisar as interações entre tornozelo, joelho, quadril, pelve e tronco durante a corrida. Três modificações de técnica de corrida foram abordadas: com aterrissagem iniciada com o antepé, com aumento de 10% na taxa de passo e com aumento de flexão de tronco, além da habitual. As análises foram realizadas para o ciclo completo (apoio e balanço) e com separação da fase de apoio, e revelaram que essas duas estratégias de processamento são complementares. Comparando as influências proximal e distal, os procedimentos sugeriram uma predominância das interações proximal a distal, mostrando uma origem central de movimentos. Dessa forma, destaca-se a relevância em controlar e fortalecer tronco e quadril para a minimização de lesões. Considerando os resultados e a oportunidade de configuração, o software pode ser empregado para estudar outras articulações e aplicações, bem como evoluir para um sistema automatizado de apoio à decisão. / Computer programs have supported the study of biomedical systems in which a considerable amount of data is employed. In biomechanics, analysis of influences between joints can improve the knowledge of the Running-Related-Injuries (RRI) associated to overuse during running activity. Understanding the patterns of interaction among anatomical joints during movement can contribute to the improvement of training, rehabilitation and injury prevention programs. In this work, a customized software was developed to implement Partial Directed Coherence (PDC), an approach in the frequency domain of Granger Causality (GC), adapted to the physical therapy specificities. With independent and standardized inputs, modularization and parameterization, the routines investigated the interaction direction between different channels, logging and saving intermediate files. Separated in the three anatomical planes, sagittal, frontal and transverse, kinematic data were employed to analyze the interactions between ankle, knee, hip, pelvis and trunk during running. Three running technique modifications were addressed: forefoot strike landing pattern, increasing 10% of the step rate and increasing trunk flexion, in addition to usual running. The analyzes were performed for the complete cycle (stance and swing) and with separation of the stance phase, and revealed that these two processing strategies are complementary. Comparing proximal and distal influences, procedures suggested a predominance of proximal to distal interactions, showing a central origin of movements. In this way, the importance of controlling and strengthening trunk and hip to minimize injuries is highlighted. Considering the results and the processing configuration opportunity, the software can be employed to study other joints and applications, as well as evolve to an automated decision support system.

Les interactions protéine-protéine (IPPs), complexes et dynamiques, sont le cœur des réseaux protéiques cellulaires. Au niveau des synapses excitatrices, la densité post-synaptique (PSD) est un exemple typique de réseau protéique dont la structure et la composition à l’échelle nanoscopique détermine la fonction cellulaire. Ainsi, la régulation dynamique de la composition de la PSD et des mouvements des récepteurs au glutamate dans ou hors de la PSD constitue la base des théories moléculaires actuelles sur l’apprentissage et la mémoire. Dans ce contexte, durant ma thèse, j’ai étudié une classe d’IPPs faisant intervenir les domaines PDZ. En effet, durant ces dernières années, de nombreuses études ont démontré l’implication de ces interactions impliquant les domaines PDZ de la famille de PSD95 dans le ciblage synaptique et l’ancrage des récepteurs au glutamate. Cependant, en partie dû au manque d’outils adaptés, les mécanismes moléculaires sous-jacents qui contrôlent de façon dynamique leur rétention à la synapse restent mal compris. Dans le but d’étudier ces interactions impliquant des domaines PDZ, j’ai développé plusieurs stratégies de sélection par phage display basées sur l’utilisation du dixième domaine de type III de la fibronectine humaine (10Fn3) dans le but de cibler les motifs d’interaction aux domaines PDZ des récepteurs (Stargazin pour les rAMPA et GluN2A pour les rNMDA) ou les domaines PDZ eux-mêmes. En utilisant une approche multidisciplinaire, mes objectifs principaux ont été de concevoir de petits anticorps synthétiques qui nous permettront de rompre ou de stabiliser spécifiquement ces complexes protéiques, ainsi que d’observer les interactions endogènes. / Complex and dynamic protein-protein interactions are the core of protein-based networks in cells. At excitatory synapses, the postsynaptic density (PSD) is a typical example of protein-based network whose nanoscale structure and composition determines the cellular function. For instance, the dynamic regulation of PSD composition and glutamate receptors movements into or out of the PSD are the base of current molecular theories of learning and memory. In this context, during my PhD, I focused on a class of protein-protein interactions mediated by PDZ domains. Indeed, over the last decade, numerous studies have shown the critical implication of PDZ domain-mediated interactions from the PSD95 scaffolding protein family in the synaptic targeting and anchoring of glutamate receptors. However, in part due to the lack of adapted tools, the molecular mechanisms that dynamically govern their respective synaptic retention remain poorly understood. In order to investigate these PDZ domain-mediated interactions, I developed several selection strategies by phage-display based on the fibronectin type III (FN3) scaffold in order to either target the PDZ domain-binding motifs of the receptors complexes (e.g., stargazin for AMPARs and GluN2A for NMDARs) or the PDZ domains themselves. Using a multidisciplinary approach, my main objectives were to engineer small synthetic antibodies that will allow us to acutely and specifically disrupt or stabilize these protein complexes, as well as monitor endogenous interactions.

La Chimie Verte s’engage entre autres à mettre au point des procédés plus respectueux de l’environnement et à émanciper de la pétrochimie les filières industrielles de fabrication de produits. Dans ce contexte, les enzymes représentent des alternatives de choix pour réaliser des réactions de synthèse de molécules écoresponsable à partir de la biomasse végétale. - L’endoglycocéramidase II de Rhodoccoccus sp. M-777, une glycoside-hydrolase, a été la cible d’un travail d'ingénierie du site actif afin de réorienter son activité hydrolytique vers la synthèse de polyglucosides d’alkyles, de potentiels biosurfactants. Une transglycosylase permettant d’atteindre des rendements de production de plus de 70% a été obtenue. La modélisation de la mutation permet de proposer des pistes sur les raisons de cette inversion du ratio hydrolyse/transglycosylation.- Une stratégie d'évolution dirigée a été appliquée à la féruloyle-estérase A d’Aspergillus niger pour la rendre plus résistante aux chocs thermiques et à la présence de solvants, deux propriétés requises pour utiliser cette enzyme pour des réactions de transfert dans des conditions thermodynamiquement favorables. Un catalogue d’enzymes améliorées, pour les deux propriétés, a été obtenu. L'accumulation de ces connaissances permettra de pouvoir plus efficacement rationaliser le design de biocatalyseur pour la synthèse de molécules, en accord avec les attentes de la chimie verte. / Green chemistry promotes the development of more environmentally friendly processes and the ending of polluting petrochemical industries by promoting the use of renewable resources. In this context, enzymes represent interesting alternatives catalysts for chemical transformations. Notably, they constitute tools of choice for synthesis of organic molecules from plant biomass.- Endoglycoceramidase II from Rhodococcus sp. M-777, a retaining glycoside hydrolase, was subjected to active-site remodelling in order to reorient its activity towards the synthesis of alkyl-polyglucosides, molecules with potential biosurfactant properties. Thus, an efficient transglycosylase able to reach production yield of more than 70% of alkyl-cellobiosides was obtained. A modelling study help to identify the determinants of this complete reversion of the transfer / hydrolysis ratio.- A directed evolution strategy was applied to Aspergillus niger feruloyl-esterase A, in order to make it more resistant to heat shocks and to the presence of solvents, two prerequisites to use this enzyme for transfer reactions under thermodynamically favourable conditions. This led to the establishment of a catalog of optimized enzymes for their thermostability, their solvent resistance, or both properties.These results will pave the way towards a more efficient way to rationally design biocatalysts meeting the expectations of green chemistry.