The Prion protein (PrP) is a cell surface glycoprotein that has been directly implicated in the pathogenesis of a range of neurological disorders referred to as the transmissible spongiform encephalopathies (TSE’s). The protein has been shown to bind copper within its unstructured N-terminus but the affinity and stoichiometry of the association is a matter of some debate. In addition, the functional significance of this copper binding has yet to be elucidated. This study aimed to determine accurate metal binding parameters for PrP through the use of calorimetry and to provide insight into the potential redox implications of metal once bound. A method of analysis for complex binding to proteins is thoroughly assessed and found to be suitable. The study also aimed to qualify the involvement of metals in the proteins remarkable ability to survive in the environment. This study confirms that PrP binds copper with an affinity relative to the amount of copper available to the protein. A high nanomolar affinity is reported within two regions on the protein, the octarepeat and the 5th site. Binding within the octarepeat region is found to be highest at low copper concentrations, reducing to micromolar affinity when copper levels exceed equivalents of 1. There is also strong evidence of a complex and cooperative binding mechanism. The 5th site also displays high nanomolar affinity for a single atom of copper. These two regions on the protein also interact in the coordination of copper (II). The copper bound protein is highly redox active and is capable of fully reversible cycling of electrons that are dependent mainly on the octarepeat. The protein does bind other divalent cations but none appear to be physiologically relevant considering the amount of these free metal ions in the body. When adsorbed to model clays, PrP is able to survive for long periods at room temperature. This longevity is increased significantly by the presence of metals in the soil, especially manganese. These data provide confirmation of the precise parameters of divalent cation binding to PrP. It also confirms that the copper bound protein is capable of a physiological redox role.

The goal of this work is to design and fabricate an autonomous acoustic recording system which is low cost, reconfigurable and portable for acoustic research. The system consists of four components, including a compact single-board computer (Celeron 1G), a 12-bit A/D converter (PCM-3718HO), a 20/40 dB gain amplifier and an useful power management circuit board. C and LabVIEW programs with multiple threads are developed to control the I/O's, digitize the underwater acoustic signals and stream data to the hard disk continuously. Currently, the throughput of data stream attains to 83 kHz. It were tested by six significant experiments to show its feasibility. At first, taking the function generator as signal source to verify no distortion caused by the system. Two ITC-6050C hydrophones are connected to the system as the receivers. The system was tested with experimental propeller producing broadband noise to calibrate the hydrophones in the experimental sink. To verify their performance, the receivers also work with the calibrated one B & K 8104 for reference. In order to understand the influence of self-noise, the system is planned to record known signal producing by hydrophone CTG-0708. By repeating similar calibration experiment, the system operates with an autonomous recording system Bioacoustic probe to understand what deficiency we should improve afterward. Further, a field test was conducted in the designated natural laboratory of shallow bay area. Post-processing of the data resulted in range-frequency plot shows that the interference pattern reported in the literature was captured. According to above-mentioned tesing, the system can be applied to underwater acoustics. This system, connected with multiple hydrophones, will be applied in the research for target detection, the harbor protection and so on.

Orientador: Marcos Roberto de Mattos Fontes / Resumo: A comunicação entre o núcleo celular e o citoplasma acontece através de mecanismos de transporte que permitem a passagem de moléculas por poros presentes no envoltório nuclear. Na Via Clássica de Importação Nuclear, a proteína Importina-α (Impα) atua na identificação das proteínas a serem transportadas ao núcleo a partir do reconhecimento de sequências de localização nuclear (NLS). Os primeiros estudos de caracterização estrutural da Impα de Neurospora crassa (NcImpα) e a sua estrutura cristalográfica mostraram a presença de regiões que podem estar relacionadas a especificidades da proteína NcImpα no reconhecimento de NLSs de proteínas de fungos. Além disso, foram reconhecidos prováveis NLSs em proteínas reguladoras do metabolismo de carbono e nitrogênio em fungos. O objetivo desse trabalho é identificar e caracterizar o modo de interação de NLSs de proteínas fúngicas com a NcImpα e verificar possíveis especificidades da proteína NcImpα no reconhecimento de NLSs. Os potenciais peptídeos NLS de proteínas dos fungos filamentosos N. crassa e Aspergillus nidulans foram submetidos a experimentos de calorimetria de titulação isotérmica (ITC) com a proteína NcImpα, para calcular a afinidade entre as moléculas. Dentre os peptídeos testados, as sequências correspondentes ao potenciais NLS dos fatores de transcrição PAC-3, NIT-2, FLB-3, VOSA e VEA, apresentaram elevada afinidade com a NcImpα, conforme indicado pelos valores de Kd obtidos. Quando submetidos a experimentos de importação ... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The communication between the cell nucleus and the cytoplasm happens through transport mechanisms that allow the passage of molecules through pores present in the nuclear envelope. In the classical nuclear import pathway, the protein Importin-α (Impα) acts in the identification of the proteins to be transported to the nucleus from the recognition of nuclear localization sequences (NLS). The first structural characterization studies of N. crassa Impα (NcImpα) and its crystallographic structure showed the presence of regions that may be related to protein specificities in the recognition of fungal NLSs. In addition, NLSs were recognized in proteins related to fungal metabolism. The objective of this work is to identify NLSs in fungal proteins and observe the specificities of the NcImpα protein. Potential NLS peptides of N. crassa were subjected to isothermal titration calorimetry (ITC) experiments with NcImpα to verify and calculate the affinity of the complexes. Among the peptides tested, the sequences corresponding to the potentials NLS of the transcription factors PAC-3, NIT-2, FLB-3, VOSA and VEA, showed high affinity with NcImpα, as indicated by the Kd values obtained. When subjected to functional experiments with HeLa cells, the peptide NIT2-NLS were efficiently transported into the cell nucleus. Crystallization tests were performed to elucidate the structure of the complexes Impα/NLS peptides. A set of data from the NcImpα/ NIT2NLS complex was collected, with 99.71% comp... (Complete abstract click electronic access below) / Doutor

Copper is an essential element for all living organisms. However, due to its low redox potential it can be involved in the production of reactive oxygen species; where excess amounts of copper can be exceptionally toxic.1 In humans, malfunctions in copper metabolism are linked to diseases such as Menkes syndrome, Wilson’s disease, prion disease, and Alzheimer’s disease.2 Maintenance of copper homeostasis requires a number of proteins, such as copper transporters and chaperones to deliver copper to the correct protein while limiting free copper in the cell.3 Therefore, understanding the thermodynamics of copper(II) coordination in proteins is critical to our understanding of copper homeostasis. Herein we report human carbonic anhydrase II contains a novel copper binding site with picomolar affinity.4 A full characterization of the structure and thermodynamics associated with the coordination of both Cu atoms into their respective sites is discussed. Techniques including paramagnetic nuclear magnetic resonance spectroscopy (NMR), and x-ray absorption spectroscopy (XAS) techniques provide insight into the high affinity CuA coordination environment. A detailed characterization of this high affinity binding site and related peptide-bound model complexes are included, with the results providing insights into the chemistry and physiological impact of copper binding in human carbonic anhydrase II.

Polymers having many ionizable groups in their molecular structure are called Polyelectrolytes. They are extensively used in industries like papermaking, food processing, medicine and pharmaceuticals, water purification, oil field exploration, cosmetic formulation etc. In spite of wide applicability its current status of knowledge is precursory due complexity of their behavior in solution as well as at interface. Solution properties of polyelectrolytes are extensively studied in last 40 years to understand their behavior. The activity is important thermodynamic property. From activity we can get most of thermodynamic properties like interaction parameter, free energy of dilution of the polyelectrolyte, degree of dissociation of polyelectrolytes etc.Several models of Polyelectrolytes thermodynamics have been proposed. Two general approaches have been used to model Polyelectrolytes thermodynamics, spherical and cylindrical (chain) models. Two of the successful models to explain and predict commonly measured properties of polyelectrolytes such as osmotic coefficient and counterion activity coefficients have been proposed by Manning and Oosawa. Most of these models are applicable at infinite dilution only may be due to weak inter chain interactions. An Excess Gibb’s free energy model can predict properties at finite concentrations of polyelectrolytes, which is combination of Manning model and Local composition model.Vapor pressure osmometry and isothermal Titration Microcalorimetry are experimental methods to determine the thermodynamic properties of polymer solvent system. Osmometry helps to understand the thermodynamics of polymer solutions as it determines the value of osmotic pressure, which in turn gives the value of thermodynamic parameters. Isothermal Titration Microcalorimetry gives the heat of dilution directly from which we can calculate activity of the solution.The osmotic coefficient and activity of water in aqueous NaPSS solution are found out using Vapor pressure osmometry and Isothermal titration calorimeter at different temperatures. The results are compared with result obtained by an excess Gibb’s free energy model. Measured data show good agreement with available literature data at that temperature.

Les télomères sont les extrémités des chromosomes linéaires des eucaryotes.Ils sont constitués de répétitions en tandem d'un motif court riche enguanine, et liés par des protéines spécifiques. Chez les vertébrés cesprotéines forment un complexe appelé le shelterin et dont l'intégrité estcritique pour assurer la réplication correcte des extrémités deschromosomes, et pour les protéger contre une prise en charge illicite parles voies de réparation des cassures double-brin de l'ADN. Des dysfonctionsdes télomères engendrent une instabilité du génome qui peut conduire à lasénescence ou au cancer. Les télomères représentent une région subnucléaireoù les protéines du shelterin sont fortement enrichies, ce qui permetl'implication dans les fonctions biologiques d'interactions de basseaffinité. Parmi les protéines du shelterin, la protéine de liaison auxrépétitions télomériques TRF2 et son partenaire constitutif RAP1 sont lesfacteurs majeurs responsables de la protection des extrémités. Nous avonsétudié en détails l'assemblage du complexe TRF2/RAP1 par des approchesintégrées de biologie structurale, de biophysique et de biochimie.Nous avons montré que cet assemblage s'accompagne d'importants ajustementsde conformation des deux protéines, et implique une interaction de basseaffinité qui engage de grandes régions des deux protéines et affecte leurspropriétés d'interactions. / Telomeres are the ends of eukaryotic linear chromosomes. They are made oftandem repeats of a short guanine-rich motif and bound by specific proteins.In vertebrates, these proteins form a complex called shelterin, theintegrity of which is critical to ensure proper replication of chromosomeends and to protect them against illicit targeting by DNA double-strandbreak repair pathways. Telomere dysfunctions lead to genome instability,which can ultimately cause senescence or cancer. Telomeres are a subnuclearregion in which shelterin proteins are highly enriched, enhancing lowaffinity interactions of important biological function. Among shelterinproteins, telomeric repeat-binding protein TRF2 and its constitutive partnerRAP1 are the main factors responsible for end protection. We studied indetails the assembly of TRF2/RAP1 complex by means of integrated structural,biophysical and biochemical approaches. We showed that this assemblydisplays important conformational adjustments of both proteins, and involvesa low affinity interaction engaging large regions in both proteins whichaffects their interaction properties.

Este trabalho tem por objetivo o estudo de algumas propriedades físicas do mineral natural sodalita, de origem brasileira (Itabuna, BA), cuja fórmula ideal é Na8Al6Si6O24Cl2. Para isso foram feitas medidas de ressonância paramagnética de elétrons, absorção óptica, ressonância nuclear magnética, corrente iônica termoestimulada e condutividade elétrica DC em amostras naturais tratadas termicamente e irradiadas com raios-X. Foram detectados três centros paramagnéticos: uma linha isotrópica com g= 2.011, relacionada com a cor azul da amostra natural, isto é, bandas de absorção óptica a 600nm e 645nm. Um conjunto de treze linhas de interação hiperfina, com g= 2.001 e A= 3.5 Gauss, relacionado com um centro de dipolo elétrico responsável por dois picos de relaxação dielétrica a 19.90K, e a 49.3&#176 K com energias de ativação de 30 meV e 121 meV respectivamente. Um conjunto de treze linhas de interação hiperfina com g= 2.001 e A= 32.5 Gauss, relacionado com uma banda de absorção óptica a 530nm / The aim of this work is the study of some physical properties of natural blue brazilian sodalite (Itabuna, BA), whose ideal formula is Na8Al6Si6O24Cl2. For this purpose, we made use of electron paramagnetic resonance, nuclear magnetic resonance, ionic thermocurrent, optical absorption and electrical conductivity techiniques in natural, bleached and irradiated samples. We have detected three paramagnetic centers: a) na isotropic line with g= 2.011, related to the blue color of natural samples, that is, with the optical absorption bands at 600nm and 645nm; b) a set of thirteen lines of hyperfine interaction with g= 2.001 and A= 3.5 Gauss, related to na electric dipole Center responsible for two bands of dieletric relaxation at 19.9&#176K and 49.3&#176K, with activation energy of 30 meV and 121 meV, respectively, c) we have also detected an F center with a EPR spectrum composed of thirteen isotropic lines of hyperfine interaction with g= 2.001 and A=32.5 Gauss, related to the Pink color

Neste trabalho realizou-se o estudo estrutural e termodinâmico de sistemas micelares em solução formados por surfactantes de diferentes classes, tais como SDS (aniônico), TTAB (catiônico), DM (não-iônico) e SB310 (zwiteriônico). A abordagem experimental proposta neste trabalho consiste do estudo simultâneo de aspectos estruturais e termodinâmicos. Para o estudo estrutural utilizou-se as técnicas de raios X à baixo ângulo (SAXS) e Condutimetria, e pelo ajuste dos dados experimentais pode-se determinar parâmetros como a anisotropia de forma, número de agregação, grau de ionização, bem como a concentração micelar crítica das micelas em solução. O comportamento da função de distribuição de pares de distância permitiu concluir que todas as micelas formam estruturas do tipo core-shell, porém diferem em tamanho dependendo da composição da micela. O buffer usado é possivelmente o responsável por causar a diminuição da concentração micelar crítica e aumento do grau de ionização (para surfactantes iônicos) quando comparados os valores obtidos neste trabalho com aqueles da literatura, que em geral usa água como solvente. Para o estudo termodinâmico utilizou-se a técnica de calorimetria de titulação isotérmica (ITC) e pelo ajuste dos dados experimentais pode-se determinar os valores de entalpia micelar e concentração micelar crítica. A combinação dos resultados obtidos das técnicas de ITC e Condutimetria permitiu calcular o valor energia livre de Gibbs e a entropia de micelização. A partir da determinação dos parâmetros descritos acima, pode-se caracterizar a estrutura e aspectos termodinâmicos de micelas em solução. A metodologia apresentada neste manuscrito consegue fornecer parâmetros independentes e ao mesmo tempo complementares, o que acaba permitindo um estudo correlacionado e confiável. / In this work was carried out the study of the structural and thermodynamic of micelar systems in solution formed by different classes of surfactants such as SDS (anionic) TTAB (cationic), DM (non-ionic) and SB310 (zwitterionic). The experimental approach proposed in this manuscript consists of the simultaneous study of structural and thermodynamic aspects. For structural studies it was utilized the techniques of Small Angle X ray Scattering (SAXS) and Conductometry, and by fitting the experimental data it was possible to evaluate parameters such as the anisotropy, aggregation number, degree of ionization, and the critical micelle concentration of the micelles in solution. From the analysis of the behavior of the pair distance distribution function we concluded that the micelles form \"core-shell\" structures, but differ in size depending on the composition of the micelle. The buffer used is possibly responsible for causing the decrease of critical micelle concentration and the increase of the degree of ionization (for ionic surfactants) when values obtained in this study are compared to those ones in the literature, which usually uses water as a solvent. For the thermodynamic study we have used the technique of Isothermal Titration Calorimetry (ITC) and by fitting the experimental data we could determine the enthalpy values and critical micelle concentration. The combination of the results of ITC and Conductometry techniques allowed us to evaluate the value of Gibbs free energy and entropy of micellization. From the determination of the parameters described above we could characterize the structure and thermodynamic aspects of the micelles in solution. The methodology presented in this work can provide independent and complementary parameters, which ends up allowing a correlated and reliable study.

Este trabalho tem por objetivo o estudo de algumas propriedades físicas do mineral natural sodalita, de origem brasileira (Itabuna, BA), cuja fórmula ideal é Na8Al6Si6O24Cl2. Para isso foram feitas medidas de ressonância paramagnética de elétrons, absorção óptica, ressonância nuclear magnética, corrente iônica termoestimulada e condutividade elétrica DC em amostras naturais tratadas termicamente e irradiadas com raios-X. Foram detectados três centros paramagnéticos: uma linha isotrópica com g= 2.011, relacionada com a cor azul da amostra natural, isto é, bandas de absorção óptica a 600nm e 645nm. Um conjunto de treze linhas de interação hiperfina, com g= 2.001 e A= 3.5 Gauss, relacionado com um centro de dipolo elétrico responsável por dois picos de relaxação dielétrica a 19.90K, e a 49.3&#176 K com energias de ativação de 30 meV e 121 meV respectivamente. Um conjunto de treze linhas de interação hiperfina com g= 2.001 e A= 32.5 Gauss, relacionado com uma banda de absorção óptica a 530nm / The aim of this work is the study of some physical properties of natural blue brazilian sodalite (Itabuna, BA), whose ideal formula is Na8Al6Si6O24Cl2. For this purpose, we made use of electron paramagnetic resonance, nuclear magnetic resonance, ionic thermocurrent, optical absorption and electrical conductivity techiniques in natural, bleached and irradiated samples. We have detected three paramagnetic centers: a) na isotropic line with g= 2.011, related to the blue color of natural samples, that is, with the optical absorption bands at 600nm and 645nm; b) a set of thirteen lines of hyperfine interaction with g= 2.001 and A= 3.5 Gauss, related to na electric dipole Center responsible for two bands of dieletric relaxation at 19.9&#176K and 49.3&#176K, with activation energy of 30 meV and 121 meV, respectively, c) we have also detected an F center with a EPR spectrum composed of thirteen isotropic lines of hyperfine interaction with g= 2.001 and A=32.5 Gauss, related to the Pink color

Neste trabalho realizou-se o estudo estrutural e termodinâmico de sistemas micelares em solução formados por surfactantes de diferentes classes, tais como SDS (aniônico), TTAB (catiônico), DM (não-iônico) e SB310 (zwiteriônico). A abordagem experimental proposta neste trabalho consiste do estudo simultâneo de aspectos estruturais e termodinâmicos. Para o estudo estrutural utilizou-se as técnicas de raios X à baixo ângulo (SAXS) e Condutimetria, e pelo ajuste dos dados experimentais pode-se determinar parâmetros como a anisotropia de forma, número de agregação, grau de ionização, bem como a concentração micelar crítica das micelas em solução. O comportamento da função de distribuição de pares de distância permitiu concluir que todas as micelas formam estruturas do tipo core-shell, porém diferem em tamanho dependendo da composição da micela. O buffer usado é possivelmente o responsável por causar a diminuição da concentração micelar crítica e aumento do grau de ionização (para surfactantes iônicos) quando comparados os valores obtidos neste trabalho com aqueles da literatura, que em geral usa água como solvente. Para o estudo termodinâmico utilizou-se a técnica de calorimetria de titulação isotérmica (ITC) e pelo ajuste dos dados experimentais pode-se determinar os valores de entalpia micelar e concentração micelar crítica. A combinação dos resultados obtidos das técnicas de ITC e Condutimetria permitiu calcular o valor energia livre de Gibbs e a entropia de micelização. A partir da determinação dos parâmetros descritos acima, pode-se caracterizar a estrutura e aspectos termodinâmicos de micelas em solução. A metodologia apresentada neste manuscrito consegue fornecer parâmetros independentes e ao mesmo tempo complementares, o que acaba permitindo um estudo correlacionado e confiável. / In this work was carried out the study of the structural and thermodynamic of micelar systems in solution formed by different classes of surfactants such as SDS (anionic) TTAB (cationic), DM (non-ionic) and SB310 (zwitterionic). The experimental approach proposed in this manuscript consists of the simultaneous study of structural and thermodynamic aspects. For structural studies it was utilized the techniques of Small Angle X ray Scattering (SAXS) and Conductometry, and by fitting the experimental data it was possible to evaluate parameters such as the anisotropy, aggregation number, degree of ionization, and the critical micelle concentration of the micelles in solution. From the analysis of the behavior of the pair distance distribution function we concluded that the micelles form \"core-shell\" structures, but differ in size depending on the composition of the micelle. The buffer used is possibly responsible for causing the decrease of critical micelle concentration and the increase of the degree of ionization (for ionic surfactants) when values obtained in this study are compared to those ones in the literature, which usually uses water as a solvent. For the thermodynamic study we have used the technique of Isothermal Titration Calorimetry (ITC) and by fitting the experimental data we could determine the enthalpy values and critical micelle concentration. The combination of the results of ITC and Conductometry techniques allowed us to evaluate the value of Gibbs free energy and entropy of micellization. From the determination of the parameters described above we could characterize the structure and thermodynamic aspects of the micelles in solution. The methodology presented in this work can provide independent and complementary parameters, which ends up allowing a correlated and reliable study.