To understand and modify the secondary cell walls of plants the project group Enzyme Discovery in Hybrid Aspen for Fiber Engineering (EDEN) was founded composed of nine laboratories with funding from the European Commission. The main target of EDEN´s research is to genetically engineer fiber structure in order to produce transgenic trees with modified properties for the pulp and paper industries. In this target framework, the Populus tremula x tremuloides xyloglucan endotransglycosylase (PttXET16A) was selected for in-depth study of its transglycosylase activity catalyzing cleavage and reconnection of xyloglucan molecules, which is proposed to be involved in secondary cell wall morphogenesis. The creation of a family 16 carbohydrate active enzyme -glucanase/XET hybrids were attempted in order to design a chimeric enzyme with one or more of the following altered properties: specificity, activity, and or stability. The two enzymes, Bacillus licheniformis 1,3-1,4--glucanase and Populus tremula x tremuloides xyloglucan endotransglycosylase, are members of the same enzymatic family and have highly homologous 3-dimensional structures. However, the enzymes exhibit different activities, one a hydrolase the other a transferase; different specificities, one accepts only linear glcosydic substrates while the other branched substrates; and different stabilities. Hybrid enzyme construction represented an investigational challenge in order to understand what physical characteristics of both enzymes attribute to the specific pattern of activity and specificity observed.Removal of the 1,3-1,4--glucanase major loop resulted in a folded protein which still maintained some β-glucan hydrolase activity. However, no xyloglucan endotransglycosylase-like activity or specificity was observed. Next, point mutations of the β-sheets forming the enzymatic binding site cleft were mutated to resemble PttXET16A residues. The final chimeric protein neither exhibited XET nor β-glucanase activities. Structural analysis by X-ray crystallography revealed a major unexpected structural rearrangement providing a clear insight for further enzyme engineering. / Amb la finalitat d'entendre i modificar la paret cel·lular secundària de les plantes, es va fundar el grup Enzyme Discovery in Hibrid Aspen for Fibern Engineering (EDEN) composat per nou laboratoris amb la finançament de la Comissió Europea. El principal objectiu de la recerca del grup EDEN és enginyar genèticament l'estructura de fibres per tal de produir arbres transgènics amb propietats modificades per les indústries de la polpa i el paper.En el marc d'aquest projecte, es va seleccionar el Populus tremula x tremuloides xiloglucà endotransglicosilasa (PttXET16A) per estudiar en profunditat la seva activitat transglicosilasa catalitzant el trencament i la reconnexió de molècules de xiloglucà, el qual sembla estar involucrat en la morfogènesi de la paret cel·lular secundària. D'aquesta manera, s'intentà crear una família 16 d'híbrids de l'enzim actiu amb carbohidrats -glucanasa/XET per tal de dissenyar un enzim quimèric amb una o més de les propietats següents alterades: especificitat, activitat i/o estabilitat.Els dos enzims, Bacillus licheniformis 1,3-1,4--glucanasa i Populus tremula x tremuloides xiloglucà endotransglicosilasa, són membres de la mateixa família enzimàtica i tenen una gran homologia en les seves estructures en 3-dimensions. Tot i així, aquests enzims presenten diferents activitats, un presenta activitat hidrolasa i l'altre, transferasa; diferents especificitats, un accepta només substrats glicosílics lineals mentre l'altre, substrats ramificats; i diferents estabilitats. La construcció d'un enzim híbrid representa un repte en la investigació amb la finalitat d'entendre quines característiques físiques dels dos enzims s'atribueixen al model específic de l'activitat i especificitat observada.L'extracció del llaç més gran de l'1,3-1,4--glucanasa va resultar en l'obtenció d'una proteïna plegada que encara manté certa activitat hidrolasa del -glucà. Tot i això, no s'observà activitat o especificitat similar a la xiloglucà endotransglicosilasa. A partir d'aquí, es realitzaren mutacions puntuals a diferents punts de les fulles  que formen l'escletxa del lloc d'unió de l'enzim per assemblar-se als residus del PttXET16A. La proteïna quimèrica final tampoc presentava activitat XET ni -glucanasa. L'anàlisi de l'estructura per cristal·lografia de raigs X revelà una major reorganització estructural de l'esperada proveint el nou enzim d'un clar espai intern que obra moltes més portes a l'enginyeria de l'enzim. / Con la finalidad de entender y modificar la pared celular secundaria de las plantas, se fundó el grupo Enzyme Discovery in Hibrid Aspen for Fibern Engineering (EDEN) compuesto por nueve laboratorios con la financiación de la Comisión Europea. El principal objetivo de la búsqueda del grupo EDEN es ingeniar genéticamente la estructura de fibras para producir árboles transgénicos con propiedades modificadas para las industrias de la pulpa y el papel. En el marco de este proyecto, se seleccionó el Populus tremula x tremuloides xiloglucán endotransglicosilasa (PttXET16A) para estudiar en profundidad su actividad transglicosilasa catalizando la rotura y la reconnexión de moléculas de xiloglucán, el cual parece estar involucrado en la morfogénesis de la pared celular secundaria. De esta forma, se intentó crear una familia 16 de híbridos de la enzima activa con carbohidratos -glucanasa/XET con la finalidad de diseñar una enzima quimérica con una o más de las propiedades siguientes alteradas: especificidad, actividad y/o estabilidad. Las dos enzimas, Bacillus licheniformis 1,3-1,4--glucanasa y Populus tremula x tremuloides xiloglucà endotransglicosilasa, son miembros de la misma familia enzimática y tienen una gran homología en sus estructuras en 3-dimensiones. Aún así, estas enzimas presentan diferentes actividades, una tiene actividad hidrolasa y la otra, transferasa; diferentes especificidades, una acepta sólo sustratos glicosílicos lineales mientras la otra, sustratos ramificados; y diferentes estabilidades. La construcción de una enzima híbrida representa un reto dentro de la investigación con la finalidad de entender qué características físicas de las dos enzimas se atribuyen al modelo específico de la actividad y especificidad observada. La extracción del lazo más grande de la 1,3-1,4--glucanasa resultó en la obtención de una proteína plegada que todavía mantiene cierta actividad hidrolasa del -glucán. Aún así, no se observó actividad o especificidad similar a la xiloglucán endotransglicosilasa. A partir de este punto, se realizaron mutaciones puntuales a diferentes puntos de las hojas  que forman la brecha del lugar de unión de la enzima por asemejarse a los residuos del PttXET16A. La proteína quimérica final tampoco presentaba actividad XET ni -glucanasa. El análisis de la estructura por cristalografía de rayos X reveló una mayor reorganización estructural de la esperada proveyendo la nueva enzima de un claro espacio interno que obre muchas más puertas a la ingeniería de la enzima.

Made available in DSpace on 2015-05-14T12:14:07Z (GMT). No. of bitstreams: 1 arquivototal.pdf: 9808595 bytes, checksum: 235eec9737496afa4dfbdf1feafbc21f (MD5) Previous issue date: 2013-02-04 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / The Dark matter (DM) problem constitutes a key question at the interface among Particle Physics, Astrophysics and Cosmology. The observational data which have been accumulated in the last years point to an existence of non baryonic amount of DM. Since the Standard Model (SM) does not provide any candidate for such non-baryonic DM, the evidence of DM is a major indication for new physics beyond the SM. We will study in this work one of the most popular DM candidates, the so called WIMPs (Weakly Interacting Massive Particles) from a direct and indirect detection perspective. In order to approach the direct and indirect dection of DM in the context of Particle Physics in a more pedagogic way, we will begin our discussion talking about a minimal extension of the SM. Later we will work on the subject in a 3-3-1 model. Next, we will study the role of WIMPs in the Big Bang Nucleosynthesis. Lastly, we will look for indirect signals of DM, by looking for gamma-ray excess observed by the NASA Satelite, called Fermi-LAT, in the Galactic Center. Through an analyses of the data events observed by Fermi-LAT and some background models, we will constrain the annihilation cross section-mass relation. As a result of this PhD 9 articles have been done. Two of them are still under the publication process. / O problema da matéria escura (ME) constitui uma questão chave na interface entre física de partículas, astrofísica e cosmologia. O acúmulo de dados observacionais nos últimos anos apontam para uma enorme quantidade de ME não bariônica. Uma vez que o Modelo Padrão (MP), não fornece um candidato para este tipo de matéria, a evidência de ME é uma forte indicação de física nova, além do MP. Estudaremos neste trabalho um dos candidatos à ME mais populares, os chamados WIMPS (partículas massivas que interagem fracamente) sob o ponto de vista de detecção direta e indireta de ME. Para que possamos abordar os meios de detecção direta e indireta de ME no contexto de Física de Partículas de forma didática, iniciaremos nossa discussão apresentando uma extensão mínima do MP. Posteriormente trataremos do assunto no contexto de um modelo 3-3-1. Adiante verificaremos qual o papel da ME no cenário da Nucleossíntese Primordial. Por ´ultimo procuraremos por sinais indiretos de ME, na busca por excessos em raios gama observados pelo satélite da NASA, chamado Fermi-LAT, no centro da nossa galáxia. Através de uma análise dos eventos observados pelo Fermi-LAT e de alguns modelos de background astrofísico iremos impor vínculos com relação à massa e seção de choque de aniquilação. Ao longo desse doutorado foram publicados 9 artigos. Um destes ainda sob o processo de publicação.

Made available in DSpace on 2014-06-11T19:32:55Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-02-06Bitstream added on 2014-06-13T21:05:14Z : No. of bitstreams: 1 vasconcelos_afd_dr_rcla.pdf: 873436 bytes, checksum: e2a838b6410dbd9cdc2bc380f1261e4e (MD5) / Exopolissacarídeos do tipo β-glucanas são polímeros produzidos por uma grande variedade de microrganismos e podem possuir diferentes propriedades físicas, químicas e aspectos estruturais. Esses biopolímeros apresentam atividades biológicas interessantes (antitumor, antiviral, anticoagulante) e aplicações comerciais como produtos em alimentos, cosméticos e farmacêuticos. Entretanto, para a aplicação dessas moléculas, é necessário primeiramente o conhecimento de suas estruturas químicas. Assim, o objetivo deste trabalho foi a produção, caracterização química de quatro exopolissacarideos (EPSGRAVIOLA, EPSMANGA, EPSPINHA e EPSLARANJA) de isolados de Botryosphaeria obtidos de frutas tropicais em decomposição e crescidos em sacarose como única fonte de carbono, determinando o melhor EPS para realizar testes de atividade anticoagulante. A homogeneidade de cada EPS foi determinada por cromatografia de filtração em gel, os quais eluíram como um único pico. Hidrólise ácida total e análise por HPAEC/PAD mostrou glucose como constituinte básico. Dados de metilação e RMN de 13C indicaram que os EPSMANGA, EPSPINHA e EPSLARANJA são glucanas lineares unidas por ligações do tipo β(1®6) e o EPSGRAVIOLA é uma glucana com ligações β(1®3) e com ramificações em C-6 de resíduos glucopiranosídicos. O espectro de FT-IR mostrou uma banda em 891 cm-1, e a espectroscopia de 13C NMR mostrou que todas as ligações eram do tipo β. Estudos realizados com o corante Vermelho Congo indicaram que os EPS possuem conformação em tripla hélice. O EPSLARANJA, uma b- D-(1®6)-glucana, foi submetido a sulfatação visando induzir a atividade anticoagulante e melhorar a solubilidade da molécula em solução, importante para a atividade biológica. Espectros de FT-IR mostraram bandas em 808 and 1252 cm-1, indicando a entrada dos grupos sulfato e as análises de RMN de 13C mostraram... / Exopolysaccharides (EPS) as β-glucans are polymers produced by a great variety of microorganisms and can possess different physical and chemical properties, and structural features. These biopolymers having interesting biological activities (antitumor, anti-viral, anticoagulant), and commercial applications in foods, cosmetics and pharmaceutical products. However, for the applications of these macromolecules, it is first necessary to understand their chemical structures. Therefore the goal of that study was the production, chemical characterization and biological activity of four exopolysaccharides (EPSGRAVIOLA, EPSMANGO, EPSPINHA and EPSORANGE) obtained from Botryosphaeria strains isolated from rotting tropical fruit grown on sucrose as carbon and the best EPS was used for the anticoagulant activity The homogeneity of each EPS was determined by gel filtration chromatography, which was eluted as a single peak. Total acid hydrolysis and HPAEC/PAD analysis of each EPS yielded only glucose. Data from methylation analysis and 13C NMR spectroscopy indicated that the EPSMANGO, EPSPINHA and EPSORANGE consisted of a linear chain of (1-6)- linked glucopyranosyl residues and EPSGRAVIOLA consisted of a main chain of glucopyranosyl (1-3) linkages substituted at O-6. FTIR spectra showed one band at 891 cm-1, and 13C NMR spectroscopy showed that all glucosidic linkages were of the β-configuration. Dye-inclusion studies with Congo Red indicated that each EPS existed in a triple-helix conformational state. The EPSORANGE, a β-(1®6)- D-glucan was submitted to a sulfation to induce anticoagulant activity and also to make this EPS more soluble, which is in favor to its biological action. The FT-IR spectrum showed bands at 808 and 1252 cm-1 indicating insertion of sulfonyl groups and the 13C NMR analysis showed that the sulfonyl groups were inserted mainly in C-4 of the b(1®6)-D-glucan... (Complete abstract click electronic access below)

In the wake of the 19th century, the protestant reformation in Europe that led to the formation of mainline reformation traditions began impacting and shaping ministry in Africa, through missionary activities. But the clarion call for Africa’s renaissance was also a wakeup call for the African church to move from being consumers of the ‘imported’ theology brewed from a European perspective and take some responsibility in producing her own theology, which can be viewed and understood with an African cultural lens. If Africa must achieve the much needed renaissance, the church certainly has an indispensable role to play. But how can there be a meaningful church praxis in African polities without a solid contextualized theological foundation? Therefore in this project, I justify the need for a biblio-centric African theology by making a case for the Cameroon Baptist Convention in the Republic of Cameroon. I argue that, to be able to solidify its foundation and enhance its Christian theological praxis in the society, a Cameroon Baptist Theological Treatise is needed in the Cameroon Baptist Convention. One that would contain well delineated and defended theological tenets required of an independent Christian denomination in Africa. Central to the development of this treatise is the development of a soteriology with a cognitive theological understanding of divine predestination. To develop a correct soteriology for this purpose, a process is required, one that would need to use sound theological principles to ensure a biblically accurate and theologically sound doctrine of soteriology as the point of departure. As an evangelical systematic theological research, I attempt in this project to deconstruct biblical and theological paradigms from scriptures and reformation traditions, which the Cameroon Baptist Convention leaders could use as tools in the process of delineating and defending their own tenets on soteriology in the proposed theological treatise. It begins by gathering data from reformation traditions and scriptures on predestination and moves on to construct theories that would not only help the Cameroon Baptist Convention leaders to develop a soteriology but also be fundamental to developing other relevant doctrines needed in the proposed Cameroon Baptist Theological Treatise. / Philosophy & Systematic Theology / D. Th. (Systematic Theology)

In the wake of the 19th century, the protestant reformation in Europe that led to the formation of mainline reformation traditions began impacting and shaping ministry in Africa, through missionary activities. But the clarion call for Africa’s renaissance was also a wakeup call for the African church to move from being consumers of the ‘imported’ theology brewed from a European perspective and take some responsibility in producing her own theology, which can be viewed and understood with an African cultural lens. If Africa must achieve the much needed renaissance, the church certainly has an indispensable role to play. But how can there be a meaningful church praxis in African polities without a solid contextualized theological foundation? Therefore in this project, I justify the need for a biblio-centric African theology by making a case for the Cameroon Baptist Convention in the Republic of Cameroon. I argue that, to be able to solidify its foundation and enhance its Christian theological praxis in the society, a Cameroon Baptist Theological Treatise is needed in the Cameroon Baptist Convention. One that would contain well delineated and defended theological tenets required of an independent Christian denomination in Africa. Central to the development of this treatise is the development of a soteriology with a cognitive theological understanding of divine predestination. To develop a correct soteriology for this purpose, a process is required, one that would need to use sound theological principles to ensure a biblically accurate and theologically sound doctrine of soteriology as the point of departure. As an evangelical systematic theological research, I attempt in this project to deconstruct biblical and theological paradigms from scriptures and reformation traditions, which the Cameroon Baptist Convention leaders could use as tools in the process of delineating and defending their own tenets on soteriology in the proposed theological treatise. It begins by gathering data from reformation traditions and scriptures on predestination and moves on to construct theories that would not only help the Cameroon Baptist Convention leaders to develop a soteriology but also be fundamental to developing other relevant doctrines needed in the proposed Cameroon Baptist Theological Treatise. / Philosophy and Systematic Theology / D. Th. (Systematic Theology)

Resumo: Exopolissacarídeos do tipo β-glucanas são polímeros produzidos por uma grande variedade de microrganismos e podem possuir diferentes propriedades físicas, químicas e aspectos estruturais. Esses biopolímeros apresentam atividades biológicas interessantes (antitumor, antiviral, anticoagulante) e aplicações comerciais como produtos em alimentos, cosméticos e farmacêuticos. Entretanto, para a aplicação dessas moléculas, é necessário primeiramente o conhecimento de suas estruturas químicas. Assim, o objetivo deste trabalho foi a produção, caracterização química de quatro exopolissacarideos (EPSGRAVIOLA, EPSMANGA, EPSPINHA e EPSLARANJA) de isolados de Botryosphaeria obtidos de frutas tropicais em decomposição e crescidos em sacarose como única fonte de carbono, determinando o melhor EPS para realizar testes de atividade anticoagulante. A homogeneidade de cada EPS foi determinada por cromatografia de filtração em gel, os quais eluíram como um único pico. Hidrólise ácida total e análise por HPAEC/PAD mostrou glucose como constituinte básico. Dados de metilação e RMN de 13C indicaram que os EPSMANGA, EPSPINHA e EPSLARANJA são glucanas lineares unidas por ligações do tipo β(1®6) e o EPSGRAVIOLA é uma glucana com ligações β(1®3) e com ramificações em C-6 de resíduos glucopiranosídicos. O espectro de FT-IR mostrou uma banda em 891 cm-1, e a espectroscopia de 13C NMR mostrou que todas as ligações eram do tipo β. Estudos realizados com o corante Vermelho Congo indicaram que os EPS possuem conformação em tripla hélice. O EPSLARANJA, uma b- D-(1®6)-glucana, foi submetido a sulfatação visando induzir a atividade anticoagulante e melhorar a solubilidade da molécula em solução, importante para a atividade biológica. Espectros de FT-IR mostraram bandas em 808 and 1252 cm-1, indicando a entrada dos grupos sulfato e as análises de RMN de 13C mostraram... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Exopolysaccharides (EPS) as β-glucans are polymers produced by a great variety of microorganisms and can possess different physical and chemical properties, and structural features. These biopolymers having interesting biological activities (antitumor, anti-viral, anticoagulant), and commercial applications in foods, cosmetics and pharmaceutical products. However, for the applications of these macromolecules, it is first necessary to understand their chemical structures. Therefore the goal of that study was the production, chemical characterization and biological activity of four exopolysaccharides (EPSGRAVIOLA, EPSMANGO, EPSPINHA and EPSORANGE) obtained from Botryosphaeria strains isolated from rotting tropical fruit grown on sucrose as carbon and the best EPS was used for the anticoagulant activity The homogeneity of each EPS was determined by gel filtration chromatography, which was eluted as a single peak. Total acid hydrolysis and HPAEC/PAD analysis of each EPS yielded only glucose. Data from methylation analysis and 13C NMR spectroscopy indicated that the EPSMANGO, EPSPINHA and EPSORANGE consisted of a linear chain of (1-6)- linked glucopyranosyl residues and EPSGRAVIOLA consisted of a main chain of glucopyranosyl (1-3) linkages substituted at O-6. FTIR spectra showed one band at 891 cm-1, and 13C NMR spectroscopy showed that all glucosidic linkages were of the β-configuration. Dye-inclusion studies with Congo Red indicated that each EPS existed in a triple-helix conformational state. The EPSORANGE, a β-(1®6)- D-glucan was submitted to a sulfation to induce anticoagulant activity and also to make this EPS more soluble, which is in favor to its biological action. The FT-IR spectrum showed bands at 808 and 1252 cm-1 indicating insertion of sulfonyl groups and the 13C NMR analysis showed that the sulfonyl groups were inserted mainly in C-4 of the b(1®6)-D-glucan... (Complete abstract click electronic access below) / Orientador: Roberto da Silva / Coorientador: Maria de Lourdes Corradi da Silva / Banca: Gabriela Alves Macêdo / Banca: Maria Inês Rezende / Banca: Jonas Contiero / Banca: Eleonora Cano Carmona / Doutor

Over a century since its inception, general relativity continues to lie at the heart of some of the most researched topics in theoretical physics. It seems likely that the coveted solutions to problems like quantum gravity are to be found in an extension of general relativity, one which may only be visible in an alternate formulation of the theory.  In this thesis we consider the possibility of casting general relativity in the form of an initial value problem where spacetime is seen as the evolution of space. This evolution is shown to be constrained and of Hamiltonian type.  Not all spacetimes are physically acceptable. To be compatible with particle physics, one would like spacetime to accommodate fermions. Here we can take comfort in Geroch’s theorem, which implies that any spacetime that admits a Hamiltonian formulation automatically supports the existence of fermions. We review the elements that go into the proof of this theorem. / Allmän relativitetsteori har i över hundra år legat i teoretiska fysikens framkant. Det är möjligt att lösningarna på öppna problem som kvantiseringen av gravitation går att finna i en utvidgning av allmän relativitetsteori – och kanske uppenbarar sig denna utvidgning bara ur en alternativ formulering av teorin. I den här uppsatsen formuleras allmän relativitetsteori och dess Einsteinekvationer som ett begynnelsevärdesproblem, genom vilket rumtiden kan betraktas som rummets historia. Vi visar att rummets rörelseekvationer är Hamiltons ekvationer med tvångsvillkor.  Enligt partikelfysiken bör fermioner kunna finnas till i rumtiden. Härom kan vi åberopa Gerochs sats, enligt vilken rumtider som har en Hamiltonsk formulering också medger fermioner. Vi redogör för huvuddragen i beviset av Gerochs sats.