Synthesis and structural analysis of the N-terminal domain of the thyroid hormone-binding protein transthyretin

Transthyretin (TTR) is a 55 kDa protein responsible for the transport of thyroid hormones and retinol in human serum. Misfolded forms of the protein are implicated in the amyloid diseases familial amyloidotic polyneuropathy and senile systemic amyloidosis. Its folding properties and stabilization by ligands are of current interest due to their importance in understanding and combating these diseases. To assist in such studies we developed a method for the solid phase synthesis of the monomeric unit of a TTR analogue and its folding to form a functional 55 kDa tetramer. The monomeric unit of the protein was chemically synthesized in three parts, comprising amino acid residues 151, 5499 and 102127, and ligated using chemoselective thioether ligation chemistry. The synthetic protein was folded and assembled to a tetrameric structure in the presence of the TTRs native ligand, thyroxine, as shown by gel filtration chromatography, native gel electrophoresis, TTR antibody recognition and thyroid hormone binding. In the current study the solution structure of the first of these fragment peptides, TTR(151) is examined to determine its intrinsic propensity to form beta-sheet structure, potentially involved in amyloid fibril formation by TTR. Despite the presence of extensive beta-structure in the native form of the protein, the Nterminal fragment adopts an essentially random coil conformation in solution.

Inter-relationships between rheumatoid arthritis and periodontal disease

This review considers the considerable similarities between periodontal disease and rheumatoid arthritis (RA). While the etiology of these two diseases may differ, the underlying pathogenic mechanisms are remarkably similar and it is possible that individuals manifesting both periodontitis and RA may suffer from a unifying underlying systemic dysregulation of the inflammatory response. In light of these findings, the implications for the use of disease-modifying medications in the management of these two chronic inflammatory conditions is apparent. Further longitudinal studies and medication-based intervention studies are required to determine just how closely these two conditions are allied.

A high definition, finite difference time domain method

A high definition, finite difference time domain (HD-FDTD) method is presented in this paper. This new method allows the FDTD method to be efficiently applied over a very large frequency range including low frequencies, which are problematic for conventional FDTD methods. In the method, no alterations to the properties of either the source or the transmission media are required. The method is essentially frequency independent and has been verified against analytical solutions within the frequency range 50 Hz-1 GHz. As an example of the lower frequency range, the method has been applied to the problem of induced eddy currents in the human body resulting from the pulsed magnetic field gradients of an MRI system. The new method only requires approximately 0.3% of the source period to obtain an accurate solution. (C) 2003 Elsevier Science Inc. All rights reserved.

Investigation of candidate division TM7, a recently recognized major lineage of the domain bacteria with no known pure-culture representatives

A molecular approach was used to investigate a recently described candidate division of the domain Bacteria, TM7, currently known only from environmental 16S ribosomal DNA sequence data, A number of TM7-specific primers and probes were designed and evaluated. Fluorescence in situ hybridization (FISH) of a laboratory scale bioreactor using two independent TM7-specific probes revealed a conspicuous sheathed-filament morphotype, fortuitously enriched in the reactor. Morphologically, the filament matched the description of the Eikelboom morphotype 0041-0675 widely associated with bulking problems in activated-sludge wastewater treatment systems. Transmission electron microscopy of the bioreactor sludge demonstrated that the sheathed-filament morphotype had a typical gram-positive cell envelope ultrastructure. Therefore, TM7 is only the third bacterial lineage recognized to have gram-positive representatives. TM7-specific FISH analysis of two full-scale wastewater treatment plant sludges, including the one used to seed the laboratory scale reactor, indicated the presence of a number of morphotypes, including sheathed filaments. TM7-specific PCR clone libraries prepared from the two full-scale sludges yielded 23 novel TM7 sequences. Three subdivisions could be defined based on these data and publicly available sequences. Environmental sequence data and TM7-specific FISH analysis indicate that members of the TM7 division are present in a variety of terrestrial, aquatic, and clinical habitats. A highly atypical base substitution (Escherichia coli position 912; C to U) for bacterial 16S rRNAs was present in almost all TM7 sequences, suggesting that TM7 bacteria, like Archaea, may be streptomycin resistant at the ribosome level.

Autosomal dominant hypocalcemia: A novel activating mutation (E604K) in the cysteine-rich domain of the calcium-sensing receptor

We report a novel activating mutation (E604K) of the calcium-sensing receptor in a family with autosomal dominant hypocalcemia. Whereas all affected individuals exhibited marked hypocalcemia, some cases with untreated hypocalcemia exhibited seizures in infancy, whereas others were largely asymptomatic from birth into adulthood. The missense mutation E604K (G2182A, GenBank accession no. U20759), which affects an amino acid residue in the C terminus of the cysteine-rich domain of the extracellular head, co-segregated with hypocalcemia in all seven individuals for whom DNA was available. Two unaffected, normocalcemic members of the family did not exhibit the mutation. The molecular impact of the mutation on two key components of the signaling response was assessed in HEK-293 cells transiently transfected with cDNA corresponding to either the wild-type calcium-sensing receptor or the E604K mutation derived by site-directed mutagenesis. There was a significant leftward shift in the concentration response curves for the effects of extracellular Ca2+ on both intracellular Ca2+ mobilization (determined by aequorin luminescence) and MAPK activity (determined by luciferase expression). The C terminus of the cysteine-rich domain of the extracellular head may normally act to suppress receptor activity in the presence of low extracellular Ca2+ concentrations.

Review of time-domain polarization measurements for assessing insulation condition in aged transformers

This paper presents a review of the time-domain polarization measurement techniques for the condition assessment of aged transformer insulation. The polarization process is first described with appropriate dielectric response theories and then commonly used polarization methods are described with special emphasis on the most widely used return voltage(rv) measurement. Most recent emphasis has been directed to techniques of determining moisture content of insulation indirectly by measuring rv parameters. The major difficulty still lies with the accurate interpretation of return voltage results. This paper investigates different thoughts regarding the interpretation of rv results for different moisture and ageing conditions. Other time domain polarization measurement techniques and their results are also presented in this paper.

GRIP domain-mediated targeting of two new coiled-coil proteins, GCC88 and GCC185, to subcompartments of the trans-Golgi network

The GRIP domain is a targeting sequence found in a family of coiled-coil peripheral Golgi proteins. Previously we demonstrated that the GRIP domain of p230/golgin245 is specifically recruited to tubulovesicular structures of the traps-Golgi network (TGN). Here we have characterized two novel Golgi proteins with functional GRIP domains, designated GCC88 and GCC185. GCC88 cDNA encodes a protein of 88 kDa, and GCC185 cDNA encodes a protein of 185 kDa. Both molecules are brefeldin A-sensitive peripheral membrane proteins and are predicted to have extensive coiled-coil regions with the GRIP domain at the C terminus. By immunofluorescence and immunoelectron microscopy GCC88 and GCC185, and the GRIP protein golgin97, are all localized to the TGN of Hela cells. Overexpression of full-length GCC88 leads to the formation of large electron dense structures that extend from the traps-Golgi. These de novo structures contain GCC88 and co-stain for the TGN markers syntaxin 6 and TGN38 but not for alpha2,6-sialyltransferase, beta-COP, or cis-Golgi GM130. The formation of these abnormal structures requires the N-terminal domain of GCC88. TGN38, which recycles between the TGN and plasma membrane, was transported into and out of the GCC88 decorated structures. These data introduce two new GRIP domain proteins and implicate a role for GCC88 in the organization of a specific TGN subcompartment involved with membrane transport.

The AF-1 Domain of the Orphan Nuclear Receptor NOR-1 Mediates Trans-activation, Coactivator Recruitment, and Activation by the Purine Anti-metabolite 6-Mercaptopurine

NOR-1/NR4A3 is an orphan member of the nuclear hormone receptor superfamily. NOR-1 and its close relatives Nurr1 and Nur77 are members of the NR4A subgroup of nuclear receptors. Members of the NR4A subgroup are induced through multiple signal transduction pathways. They have been implicated in cell proliferation, differentiation, T-cell apoptosis, chondrosarcomas, neurological disorders, inflammation, and atherogenesis. However, the mechanism of transcriptional activation, coactivator recruitment, and agonist-mediated activation remain obscure. Hence, we examined the molecular basis of NOR-1-mediated activation. We observed that NOR-1 trans-activates gene expression in a cell- and target-specific manner; moreover, it operates in an activation function (AF)-1-dependent manner. The N-terminal AF-1 domain delimited to between amino acids 1 and 112, preferentially recruits the steroid receptor coactivator (SRC). Furthermore, SRC-2 modulates the activity of the AF-1 domain but not the C-terminal ligand binding domain (LBD). Homology modeling indicated that the NOR-1 LBD was substantially different from that of hRORbeta, a closely related AF-2-dependent receptor. In particular, the hydrophobic cleft characteristic of nuclear receptors was replaced with a very hydrophilic surface with a distinct topology. This observation may account for the inability of this nuclear receptor LBD to efficiently mediate cofactor recruitment and transcriptional activation. In contrast, the N-terminal AF-1 is necessary for cofactor recruitment and can independently conscript coactivators. Finally, we demonstrate that the purine anti-metabolite 6-mercaptopurine, a widely used antineoplastic and anti-inflammatory drug, activates NOR-1 in an AF-1-dependent manner. Additional 6-mercaptopurine analogs all efficiently activated NOR-1, suggesting that the signaling pathways that modulate proliferation via inhibition of de novo purine and/or nucleic acid biosynthesis are involved in the regulation NR4A activity. We hypothesize that the NR4A subgroup mediates the genotoxic stress response and suggest that this subgroup may function as sensors that respond to genotoxicity.

Pol??ticas y pr??cticas para gestionar los conflictos de inter??s en Europa: una visi??n comparada

Este informe expone de forma sumaria los diversos enfoques y aproximaciones que nueve pa??ses de la Uni??n Europea ??? seis antiguos miembros (Reino Unido, Alemania, Francia, Italia, Espa??a y Portugal) y tres miembros recientes (Letonia, Polonia y Hungr??a) ??? est??n usando para gestionar los conflictos de inter??s en el sector p??blico. En la primera parte, se presenta el marco conceptual y las definiciones con las que poder entender de qu?? hablamos cuando hablamos de corrupci??n y de conflictos de inter??s en el sector p??blico. Posteriormente, trata de las razones existentes tras las pol??ticas de regulaci??n y tratamiento de los conflictos de inter??s y la importancia de ellas para el funcionamiento adecuado de la democracia. A continuaci??n, se explican las caracter??sticas, peculiaridades y dilemas de esta pol??tica p??blica en el contexto de las teor??as y enfoques sobre pol??ticas p??blicas. El informe tambi??n examina los puntos comunes en las estructuras, m??todos y procesos utilizados para gestionar los conflictos de inter??s en los pa??ses estudiados, adem??s de explicitar las principales diferencias en los marcos legales, medios de implantaci??n, mecanismos de evaluaci??n y medios de ejecuci??n. Por ??ltimo, incluye una visi??n cr??tica de las ventajas y desventajas relacionadas con la utilidad de los instrumentos empleados y finaliza con recomendaciones relativas a c??mo formular e implantar pol??ticas de este tipo.

Using objects and patterns to implement domain ontologies

Ontologies are becoming an important mechanism to build information systems. Nevertheless, there is still no systematic approach to support the design of such systems using tools that are common to information systems developers. In this paper, we propose an approach for deriving object frameworks from domain ontologies and then we show the application of this approach in the software process domain.

Redes de saberesfazeres, complexidade e tensões : os cotidianos do ensino médio de uma escola pública do Espírito Santo

A ciência com suas verdades passou a ser um artefato essencial nos cotidianos escolares. Esses saberes se enredam com outros saberes que despotencializados e silenciados, permanecem nos interstícios do discurso hegemônico da modernidade na escola causando tensões. Potencializar a ocorrência dessas redes nesses cotidianos pode intensificar essas tensões e assim abalar esse domínio da ciência. Os muitos caminhos possíveis de serem percorridos nas tessituras desprivilegiam quaisquer saberes e fazeres que se pretendem exclusivos. Assumindo a metáfora das redes, os praticantes da escola podem potencializar as inter-relações que ocorrem na vida. A remoção de instâncias de poder hierárquico de saberes, enredando-os e o compartilhamento solidário das práticasteoriaspráticas, não significa a diluição da autoridade. Agora a autoridade cumpre um novo papel que é de constituir uma rede de relações de autoridade partilhada. Desejamos perceber esses múltiplos sentidos e problematizar o quão potentes são ou não para a vida. Participamos juntamente com os alunos e alunas das aulas e posteriormente, os convidamos para conversasentrevistas, gravando e fotografando esses momentos. Percorremos, nos turnos matutino e noturno, todos os seus espaçostempos, nos conectando nas redes que se tecem. Optamos por estar nas turmas de ensino médio de formação geral e nas turmas de ensino médio na modalidade Educação de Jovens e Adultos. As entrevistasconversas foram feitas com três alunos ao mesmo tempo, sempre de uma mesma turma, para que se estabelecesse uma conversa e para que a produção de sentidos dos cotidianos da escola pudesse ser mais potencializada. Dos alunos entrevistados, estão entre os seus desejos mais comuns para além dos saberes científicos, as atividades esportivas, artísticas, literárias, religiosas, cursos de formação profissional, dança, canto, campanhas educativas, passeios. As relações de amizade e outros modos relacionais como entre professores e alunos, têm sido citados com grande frequência, pelos estudantes, nas entrevistasconversas. Não tem se conseguido implantar na sua totalidade os preceitos da modernidade nos escola. Há, portanto, um visível esgotamento do monopólio do comportamento científico nos cotidianos da escola pública pesquisada. São nesses espaçostempos escolares nos quais pode se dar a experiência. Apesar do discurso moderno na escola produzir mecanismos que impedem a experiência, esse controle é acompanhado de diversos mecanismos de escapes. É pela “voz” dos alunos que se apresentam as ausências da escola. A “voz” dos estudantes é a grande presença na escola e sua grande potência. Talvez esse seja o maior desperdício no contexto educacional escolar.

A formação de educadores e a travessia de cercas invisíveis de acesso/produção de conhecimentos : experiências do MST nas inter-relações com universidades brasileiras /

Este estudo sistematiza e aborda a temática da formação de educadores desenvolvida nas inter-relações entre universidades brasileiras e o Movimento dos Trabalhadores Rurais Sem Terra, especificamente nos cursos de graduação em História na Universidade Federal da Paraíba e de Engenharia Agronômica na Universidade Federal de Sergipe, no período de 2004 a 2008. Tal processo desenvolve-se em um contexto difícil e complexo da luta pela reforma agrária no Brasil, principalmente pelas transformações ocorridas nos últimos anos oriundas da ampliação das lógicas de produção do agronegócio. Esta condição leva o MST a discutir e propor uma nova concepção de reforma agrária que a designa de popular em substituição à proposta de reforma agrária clássica. Por outro lado, apresenta uma visão das universidades brasileiras e dos projetos que são construídos e implementados nos últimos anos, inclusive, observando coincidências com as políticas econômicas gerais para a sociedade. Apresenta uma concepção de formação que vem sendo construída no interior das práticas do MST que também procura as universidades para firmar convênios e desenvolver processos de escolarização/formação de seus militantes educadores. Dentre as dimensões desse processo educativo/formativo, destacam-se: vínculo permanente com os processos orgânicos; a formação como um processo ético, estético, místico que trata das atitudes/comportamentos e como um processo dialógico, crítico e articulado que contempla saberes, experiências, em uma interação que busca superar as monoculturas. Captura por intermédio da pesquisa de campo: estranhamentos, entraves, sentidos da ocupação pedagógica e coletiva, alternativas, legados que permanecem tanto no MST como na universidade e apresenta o resultado do envolvimento e atuação dos egressos de ambos os cursos na atualidade. Aponta também para desafios, possibilidades outras de enfrentar a difícil mas necessária tarefa de formar educadores, militantes capazes de coletivamente levar adiante a luta por um mundo mais justo, solidário e democrático, em que a terra e o conhecimento, juntamente com os demais bens econômicos e culturais sejam profundamente democratizados. Pretende ser uma contribuição para o debate acerca da relevância dessas inter-relações entre universidade e movimentos sociais, em que essas experiências demarcam novas possibilidades de abertura e avanços democráticos e menos elitista da universidade e novos patamares de escolarização/formação para integrantes do Movimento dos Sem Terra.

O peso das relações inter-regionais com a União Européia em relação a outras alternativas de política externa do Mercosul

No artigo examina-se a importância das relações inter-regionais entre o Mercosul e a União Européia, em particular em relação a outras alternativas de política externa.

Pope Francis and the challenges of inter-civilization diplomacy

Abstract This article addresses the role of Pope Francis in the interfaith and inter-civilization dialogue of the Holy See. Specifically, it analyses the main aspects of diplomatic relations between the Holy See and Russia, China, Islamic countries and Israel, and provides perspectives on the inter-civilization diplomacy of the Holy See during the current pontificate.