The Catharanthus roseus 16-hydroxytabersonine O-methyltransferase involved in vindoline biosynthesis /

Madagascar periwinkle (Catharanthus roseus) produces the well known and remarkably complex dimeric anticancer alkaloids vinblastine and vincristine that are derived by coupling vindoline and catharanthine monomers. This thesis describes the novel application of carborundum abrasion (CA) technique as a tool for large scale isolation of leaf epidermis enriched proteins. This technique was used to facilitate the purification to apparent homogeneity of 16-hydroxytabersonine-16-0-methyltransferse (l60MT) that catalyses the second step in the 6 step pathway that converts tabersonine into vindoline. This versatile tool was also used to harvest leaf epidermis enriched mRNAs that facilitated the molecular cloning of the 160MT. Functional expression and biochemical characterization of recombinant 160MT enzyme showed that it had a very narrow substrate specificity and high affinity for 16-hydroxytabersonine, since other closely related monoterpene indole alkaloids (MIAs) did not act as substrates. In addition to allowing the cloning of this gene, CA technique clearly showed that 160MT is predominantly expressed in Catharanthus leaf epidermis, in contrast to several other OMTs that appear to be expressed in other Catharanthus tissues. The results provide compelling evidence that most of the pathway for vindoline biosynthesis including the 0- methylation of 16-hydroxytabersonine occurs exclusively in leaf epidermis, with subsequent steps occurring in other leaf cell types. Small molecule O-methyltransferases (OMTs) (E.C. 2.1.1.6.x) catalyze the transfer of the reactive methyl group of S-adenosyl-L-methionine (SAM) to free hydroxyl groups of acceptor molecules. Plant OMTs, unlike their monomeric mammalian homologues, exist as functional homodimers. While the biological advantages for dimer fonnation with plant OMTs remain to be established, studies with OMTs from the benzylisoquinoline producing plant, Thalictrum tuberosum, showed that co-expression of 2 recombinant OMTs produced novel substrate specificities not found when each rOMT was expressed individually (Frick, Kutchan, 1999) . These results suggest that OMTs can fonn heterodimers that confer novel substrate specificities not possible with the homodimer alone. The present study describes a 160MT model based strategy attempting to modify the substrate specificity by site-specific mutagenesis. Our failure to generate altered substrate acceptance profiles in our 160MT mutants has lead us to study the biochemical properties ofhomodimers and heterodimers. Experimental evidence is provided to show that active sites found on OMT dimers function independently and that bifunctional heterodimeric OMTs may be fonned in vivo to produce a broader and more diverse range of natural products in plants.

Synthèse totale du méthyl ester de la zincophorine et étude DFT du transfert d’hydrogène diastéréosélectif impliquant des intermédiaires radicalaires vicinaux à un ester

Cet ouvrage traite d’une méthodologie pour l’induction de centres stéréogènes au sein des motifs propionates par la chimie des radicaux et de son application à la synthèse totale du méthyl ester de la zincophorine. Cet ionophore, aux propriétés biologiques intéressantes, présente plusieurs défis synthétiques dont une séquence de type polypropionate anti, anti, anti, anti difficilement accessible ainsi qu’un tétrahydropyrane trans trisubstitué. Récemment, l’intérêt renouvelé pour ces composés polyéthers, en tant qu’agents anticancéreux, accentue l’importance de stratégies versatiles permettant l’accès à ces structures ainsi qu’à leurs analogues. Depuis quelques années, notre groupe s’intéresse à la synthèse d’unités propionates acycliques par une séquence réactionnelle contrôlée uniquement par le substrat. La première étape découle d’une aldolisation de Mukaiyama entre un aldéhyde alpha-chiral et un énoxysilane tétrasubstitué portant une liaison carbone-halogène, et où l’issue stéréochimique dépend de la nature monodentate ou bidentate de l’acide de Lewis employé. La seconde réaction-clé implique la formation d’un radical tertiaire, vicinal à un ester, pouvant être réduit diastéréosélectivement en présence d’hydrure d’étain. La première section décrit l’élaboration de motifs tétrahydropyranes trisubstitués et l’induction des centres stéréogènes vicinaux par une réduction radicalaire. Nous avons révélé que l’issue diastéréosélective de la réaction de cyclisation par une iodoéthérification était dictée par le groupement gamma-méthyle des esters alpha,beta-insaturés de départ. Nous avons ensuite démontré que les produits de la réaction radicalaire anti et syn pouvaient être obtenus sélectivement à partir d’un intermédiaire commun, respectivement en prenant appui sur l’effet exocyclique ou endocyclique lors de la réduction. Par une stratégie complémentaire, nous avons révélé que le précurseur radicalaire pouvait également être obtenu par une réaction de cycloéthérification en présence d’un énoxysilane tétrasubstitué. Une étude systématique des substituants du cycle a révélé que certaines relations stéréochimiques conduisaient à une perte de sélectivité au détriment du produit anti. La seconde section concerne l’étude DFT au niveau BHandHLYP/TZVP des intermédiaires radicalaires impliqués lors du transfert d’hydrogène. Par une étude de décomposition de l’énergie d’activation, nous avons été en mesure de rationaliser l’issue diastéréosélective de la réaction sur la base des énergies de distorsion (∆Ed‡) et d’interaction (∆Eint‡) requises pour accéder à la paire d’états de transition pro-anti et pro-syn. De plus, nous avons démontré qu’une analyse NBO permettait de relativiser l’impact des interactions stéréoélectroniques. Par la suite, l’évaluation des intermédiaires radicalaires borinates et aluminates nous a permis de révéler que l’encombrement stérique de la chaîne propionate était la cause principale de la formation sélective des produits anti ou syn lors d’une réduction en présence d’un acide de Lewis. La dernière section décrit l’élaboration de la séquence polypropionate de la zincophorine, et de ses isomères, à partir du fragment tétrahydropyrane substitué. Au cours de notre étude, nous avons identifié que le nombre de sites de chélation potentiels devait être limité à trois lors de l’aldolisation en condition Cram-chélate. De plus, nous avons démontré que les différents motifs acétates sont accessibles sélectivement par l’utilisation d’un énoxysilane encombré. Par ailleurs, nous avons révélé qu’une même séquence réactionnelle pouvait être employée pour la synthèse du fragment C17–C25 de son analogue naturel CP-78,545, et avons complété la plus récente synthèse totale du méthyl ester de la zincophorine.

Liderazgo y resiliencia organizacional en la crisis financiera global del 2008–2009

Durante la crisis financiera global de 2008 muchas organizaciones y mercados financieros tuvieron que terminar sus operaciones o replantearlas debido a los choques que golpearon el bienestar de sus empresas. A pesar de esta grave situación, en la actualidad se pueden encontrar empresas que se recuperaron y salieron del terrible panorama que les presentó la crisis, incluso encontrando nuevas oportunidades de negocio y fortaleciendo su futuro. Esta capacidad que algunas organizaciones tuvieron y que permitió su salida victoriosa de la crisis se denomina resiliencia, la cual es la capacidad de sobreponerse a los efectos negativos de choques internos o externos (Briguglio, Cordina, Farrugia & Vella 2009). Por tanto en el presente trabajo se estudiará esta capacidad tanto en la organización como en los líderes para hallar factores que mejoren el desempeño de las empresas en crisis como la que ocurrió en el 2008 – 2009. Primero se realizará un estudio sobre los sucesos y el desarrollo de la crisis subprime del año 2008 para tener un entendimiento claro de sus antecedentes, desarrollo, magnitud y consecuencias. Posteriormente se realizará un estudio profundo sobre la teoría de la resiliencia organizacional y la resiliencia en el líder como individuo y los estilos de liderazgo. Finalmente teniendo un sustento teórico tanto de la crisis como del concepto de resiliencia se tomarán casos de estudio de empresas que lograron perdurar en la crisis financiera del 2008 y empresas que no lograron sobrevivir para posteriormente hallar características del líder y del liderazgo que puedan aumentar o afectar la capacidad de resiliencia de las organizaciones con el objetivo de brindar herramientas a los líderes actuales para que manejen de forma eficiente y eficaz las empresas en un mundo complejo y variable como el actual.

Cloud bands over southern Africa: seasonality, contribution to rainfall variability, and modulation by the MJO

Tropical-extratropical cloud band systems over southern Africa, known as tropical temperate troughs (TTTs), are known to contribute substantially to South African summer rainfall. This study performs a comprehensive assessment of the seasonal cycle and rainfall contribution of TTTs by using a novel object-based strategy that explicitly tracks these systems for their full life cycle. The methodology incorporates a simple assignment of station rainfall data to each event, thereby creating a database containing detailed rainfall characteristics for each TTT. This is used to explore the importance of TTTs for rain days and climatological rainfall totals in October–March. Average contributions range from 30 to 60 % with substantial spatial heterogeneity observed. TTT rainfall contributions over the Highveld and eastern escarpment are lower than expected. A short analysis of TTT rainfall variability indicates TTTs provide substantial, but not dominant, intraseasonal and interannual variability in station rainfall totals. TTTs are however responsible for a high proportion of heavy rainfall days. Of 52 extreme rainfall events in the 1979–1999 period, 30 are associated with these tropical-extratropical interactions. Cut-off lows were included in the evolution of 6 of these TTTs. The study concludes with an analysis of the question: does the Madden-Julian Oscillation influence the intensity of TTT rainfall over South Africa? Results suggest a weak but significant suppression (enhancement) of intensity during phase 1(6).

Searching for happiness across cultures

Three experiments examined the cultural relativity of emotion recognition using the visual search task. Caucasian-English and Japanese participants were required to search for an angry or happy discrepant face target against an array of competing distractor faces. Both cultural groups performed the task with displays that consisted of Caucasian and Japanese faces in order to investigate the effects of racial congruence on emotion detection performance. Under high perceptual load conditions, both cultural groups detected the happy face more efficiently than the angry face. When perceptual load was reduced such that target detection could be achieved by feature-matching, the English group continued to show a happiness advantage in search performance that was more strongly pronounced for other race faces. Japanese participants showed search time equivalence for happy and angry targets. Experiment 3 encouraged participants to adopt a perceptual based strategy for target detection by removing the term 'emotion' from the instructions. Whilst this manipulation did not alter the happiness advantage displayed by our English group, it reinstated it for our Japanese group, who showed a detection advantage for happiness only for other race faces. The results demonstrate cultural and linguistic modifiers on the perceptual saliency of the emotional signal and provide new converging evidence from cognitive psychology for the interactionist perspective on emotional expression recognition.

Regional business networks in the multinational retail sector

This article examines the network relationships of a set of large retail multinational enterprises (MNEs). We analyze under what conditions a flagship-network strategy (characterized by a network of five partners – the MNE, key suppliers, key partners, selected competitors and key organisations in the non-business infrastructure) explains the internationalisation of three retailers whose geographic scope, sectoral conditions and competitive strategies differ substantially. We explore why and when retailers will adopt a flagship strategy. The three firms are two U.K.-based multinational retailers (Tesco and The Body Shop) and a French-based global retailer (Moët Hennessy Louis Vuitton). We find evidence of strong network relationships for all three retailers, although each embraces network strategies for different reasons. Their flagship relationships depend on each retailer's strategic use of firm-specific-advantages (FSAs) and country-specific advantages (CSAs). We find that a flagship strategy can succeed in overcoming internal and/or environmental constraints to cross-border resource transfers, which are barriers to foreign direct investment (FDI). We provide recommendations on why and when to use a flagship-based strategy and which type of network partners to prioritize in order to succeed internationally.

GPS/VRS positioning using atmospheric modeling

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Uma estratégia dirigida a modelos e baseada em linguagem de descrição arquitetural para linhas de produtos de software

Model-oriented strategies have been used to facilitate products customization in the software products lines (SPL) context and to generate the source code of these derived products through variability management. Most of these strategies use an UML (Unified Modeling Language)-based model specification. Despite its wide application, the UML-based model specification has some limitations such as the fact that it is essentially graphic, presents deficiencies regarding the precise description of the system architecture semantic representation, and generates a large model, thus hampering the visualization and comprehension of the system elements. In contrast, architecture description languages (ADLs) provide graphic and textual support for the structural representation of architectural elements, their constraints and interactions. This thesis introduces ArchSPL-MDD, a model-driven strategy in which models are specified and configured by using the LightPL-ACME ADL. Such strategy is associated to a generic process with systematic activities that enable to automatically generate customized source code from the product model. ArchSPLMDD strategy integrates aspect-oriented software development (AOSD), modeldriven development (MDD) and SPL, thus enabling the explicit modeling as well as the modularization of variabilities and crosscutting concerns. The process is instantiated by the ArchSPL-MDD tool, which supports the specification of domain models (the focus of the development) in LightPL-ACME. The ArchSPL-MDD uses the Ginga Digital TV middleware as case study. In order to evaluate the efficiency, applicability, expressiveness, and complexity of the ArchSPL-MDD strategy, a controlled experiment was carried out in order to evaluate and compare the ArchSPL-MDD tool with the GingaForAll tool, which instantiates the process that is part of the GingaForAll UML-based strategy. Both tools were used for configuring the products of Ginga SPL and generating the product source code

The Leishmania amazonensis TRF ( TTAGGG repeat-binding factor) homologue binds and co-localizes with telomeres

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Foraging Behavior and Mortality of Ectatomma brunneum (Hymenoptera, Formicidae) in Simultaneous Exposure to Ant Baits and Conventional Diet in Laboratory

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

The putative telomerase reverse transcriptase component of Leishmania amazonensis: gene cloning and characterization

The Leishmania amazonensis telomerase gene was cloned by a polymerase chain reaction-based strategy using primers designed from a Leishmania major sequence that shared similarities with conserved telomerase motifs. The genes from three other species were cloned for comparative purposes. A ClustalW multiple-sequence alignment demonstrated that the Leishmania telomerases show greater homology with each other than with the proteins of other kinetoplastids and eukaryotes. Characterization experiments indicated that the putative Leishmania telomerase gene was probably in single copy and located in the largest chromosomes. A single messenger ribonucleic acid transcript was found in promastigotes. Phylogenetic analysis suggested that Leishmania telomerase might represent a liaison between the oldest and the newest branches of telomerases.

Yachting in the Eastern Caribbean: a regional overview

Includes bibliography

Impactos da restrição nutricional em cabritos Saanen dos 5 aos 15 Kg de peso corporal

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Efeitos da alimentação artificial protéica em colônias de uruçu-cinzenta (Melípona fasciculata, Smith 1858) (Apidae, Meliponini) e adaptação em casa-de-vegetaçao

O trabalho teve como objetivo desenvolver uma alimentação artificial protéica a base de extrato de soja (saburá artificial), e avaliar o seu efeito sobre a razão sexual, longevidade de operárias e desenvolvimento de colônias recém-divididas de Melipona fasciculata além de verificar a adaptação da espécie dentro de casas de vegetação. O saburá artificial aqui desenvolvido é constituído de 50g de extrato de soja, 20g de saburá fresco e 60ml de xarope de açúcar invertido (60%). Foi utilizado anilina para colorir o saburá artificial na tentativa de rastrear e verificar o consumo pelas operárias dentro das colônias. Foram utilizadas cinco colônias, das quais três receberam somente saburá e duas o saburá artificial. Não houve diferença significativa entre a produção de rainhas e operárias nos dois tratamentos e nos dois casos não houve produção de machos. As operárias que nasceram de caixas alimentadas com saburá artificial apresentaram maior longevidade e menor peso ao nascer. Estas caixas ainda iniciaram o processo de construção de células e postura mais cedo que as caixas alimentadas com saburá, contudo, suas rainhas apresentaram menor taxa de oviposição diária. O alimento a base de extrato de soja (saburá artificial) não afetou negativamente colônias recém divididas de M. fasciculata. Nos primeiros dias, as abelhas passaram a maior parte do seu tempo no topo da casa-de-vegetação tentando fugir, somente após o terceiro dia houve redução na mortalidade das operárias. Não houve diferença significativa, ao longo de cinco dias, entre a mortalidade de operárias em caixas transferidas, durante a noite e durante o dia, para dentro da casa-de-vegetação. A anilina se mostrou uma excelente ferramenta para controlar os alimentos manuseados e consumidos pela colônia.

Predictive factors of post-discharge surgical site infections among patients from a teaching hospital

Introduction Surgical site infections (SSIs) often manifest after patients are discharged and are missed by hospital-based surveillance. Methods We conducted a case-reference study nested in a prospective cohort of patients from six surgical specialties in a teaching hospital. The factors related to SSI were compared for cases identified during the hospital stay and after discharge. Results Among 3,427 patients, 222 (6.4%) acquired an SSI. In 138 of these patients, the onset of the SSI occurred after discharge. Neurological surgery and the use of steroids were independently associated with a greater likelihood of SSI diagnosis during the hospital stay. Conclusions Our results support the idea of a specialty-based strategy for post-discharge SSI surveillance.