Expression, purification and kinetic characterization of His-tagged glyceraldehyde-3-phosphate dehydrogenase from Trypanosoma cruzi

Trypanosomes are flagellated protozoa responsible for serious parasitic diseases that have been classified by the World Health Organization as tropical sicknesses of major importance. One important drug target receiving considerable attention is the enzyme glyceraldehyde-3-phosphate dehydrogenase from the protozoan parasite Trypanosoma cruzi, the causative agent of Chagas disease (T. cruzi Glyceraldehyde-3-phosphate dehydrogenase (TcGAPDH); EC 1.2.1.12). TcGAPDH is a key enzyme in the glycolytic pathway of T. cruzi and catalyzes the oxidative phosphorylation of D-glyceraldehyde-3-phosphate (G3P) to 1,3-bisphosphoglycerate (1,3-BPG) coupled to the reduction of oxidized nicotinamide adenine dinucleotide, (NAD(+)) to NADH, the reduced form. Herein, we describe the cloning of the T. cruzi gene for TcGAPDH into the pET-28a(+) vector, its expression as a tagged protein in Escherichia coli, purification and kinetic characterization. The His(6)-tagged TcGAPDH was purified by affinity chromatography. Enzyme activity assays for the recombinant His(6)-TcGAPDH were carried out spectrophotometrically to determine the kinetic parameters. The apparent Michaelis-Menten constant (K(M)(app)) determined for D-glyceraldehyde-3-phosphate and NAD(+) were 352 +/- 21 and 272 +/- 25 mu M, respectively, which were consistent with the values for the untagged enzyme reported in the literature. We have demonstrated by the use of Isothermal Titration Calorimetry (ITC) that this vector modification resulted in activity preserved for a higher period. We also report here the use of response surface methodology (RSM) to determine the region of optimal conditions for enzyme activity. A quadratic model was developed by RSM to describe the enzyme activity in terms of pH and temperature as independent variables. According to the RMS contour plots and variance analysis, the maximum enzyme activity was at 29.1 degrees C and pH 8.6. Above 37 degrees C, the enzyme activity starts to fall, which may be related to previous reports that the quaternary structure begins a process of disassembly. (C) 2010 Elsevier Inc. All rights reserved.

Fast Acquisition of (13)C NMR Spectra using the Steady-state Free Precession Sequence

In this paper, we show that the steady-state free precession sequence can be used to acquire (13)C high-resolution nuclear magnetic resonance spectra and applied to qualitative analysis. The analysis of brucine sample using this sequence with 60 degrees flip angle and time interval between pulses equal to 300 ms (acquisition time, 299.7 ms; recycle delay, 300 ms) resulted in spectrum with twofold enhancement in signal-to-noise ratio, when compared to standard (13)C sequence. This gain was better when a much shorter time interval between pulses (100 ms) was applied. The result obtained was more than fivefold enhancement in signal-to-noise ratio, equivalent to more than 20-fold reduction in total data recording time. However, this short time interval between pulses produces a spectrum with severe phase and truncation anomalies. We demonstrated that these anomalies can be minimized by applying an appropriate apodization function and plotting the spectrum in the magnitude mode.

Role of Culture in the Acquisition of CSL – From the Perspective of Intercultural Communication Competence

In the past few decades, the demands for coping with the rapid development of information communication technology, internationalization and globalization worldwide have shifted the focus of Chinese as a second language (CSL) towards intercultural communication competence in which the role of culture in the acquisition of CSL and in the pragmatic use of the language is emphasized and promoted. However, most of the present research in this academic area still remains only on a theoretical level.   In order to explore the possibilities and limitations of integrating Chinese culture and implementing intercultural communication theory into CSL education, an action research has been conducted since the beginning of 2013 to review an actual course for beginners. This paper will present the findings of the research: 1) By applying the theoretical framework of intercultural communicative competence, the findings indicated that the existing CSL course provided limited information explaining the cultural elements that are reflected in the Chinese language. 2) The findings also suggested that the cultural skills acquired in the students’ first language do influence their acquisition of CSL. This is demonstrated in the students’ written tasks such as introducing themselves and presenting other people, etc. The findings can be examples and resources for further research in this academic field.     

On insurance markets with endogenous information acquisition: A robust mechanism design approach

Neste trabalho propomos a aplicação das noções de equilíbrio da recente literatura de desenho de mecanismo robusto com aquisição de informação endógena a um problema de divisão de risco entre dois agentes. Através deste exemplo somos capazes de motivar o uso desta noção de equilíbrio, assim como discutir os efeitos da introdu ção de uma restrição de participação que seja dependente da informação. A simplicidade do modelo nos permite caracterizar a possibilidade de implementar a alocação Pareto efiente em termos do custo de aquisição da informação. Além disso, mostramos que a precisão da informação pode ter um efeito negativo sobre a implementação da alocação efi ciente. Ao final, sao dados dois exemplos específicos de situações nas quais este modelo se aplica.

Mid-auction information acquisition

This paper studies a model of a sequential auction where bidders are allowed to acquire further information about their valuations of the object in the middle of the auction. It is shown that, in any equilibrium where the distribution of the final price is atornless, a bidder's best response has a simple characterization. In particular, the optimal information acquisition point is the same, regardless of the other bidders' actions. This makes it natural to focus on symmetric, undominated equilibria, as in the Vickrey auction. An existence theorem for such a class of equilibria is presented. The paper also presents some results and numerical simulations that compare this sequential auction with the one-shot auction. 8equential auctions typically yield more expected revenue for the seller than their one-shot counterparts. 80 the possibility of mid-auction information acquisition can provide an explanation for why sequential procedures are more often adopted.

Start-up acquisition: motivations, difficulties and lesson learned

Em um esforço continuo de sobrevivência, corporações buscam meios para expandir seus negócios, seja dentro de seus mercados atuantes, ou na exploração de novos mercados. Para alcançar esse objetivo, inovações são realizadas pelos funcionários que, por meio de suas iniciativas, praticam o empreendedorismo coorporativo. São diversas formas em que o empreendedorismo corporativo se manifesta, dentre elas por meio de aquisições. Ao comprar uma outra empresa, a empresa compradora tem como motivação a busca de alguns atributos que complementem seus objetivos iniciais. Após a compra, o processo de integração entre as duas empresas é, por muitas vezes, complicado e penoso. O objetivo desse estudo é o de identificar quais são essas motivações de compra, os problemas enfrentados durante a integração entre as duas empresas, e quais são as lições aprendidas por grandes/médias corporações quando adquirem Start-ups. Muitos estudos atualmente tratam de temas relacionados à aquisição de empresas, porém, quando se trata de Start-ups, pouca literatura é encontrada. Os resultados dessa pesquisa são fruto de entrevistas com os gestores e empreendedores que participaram do processo de aquisição da SAMURAI pela Momentum e da Save-me pela Buscapé – empresas brasileiras. Os resultados são seis sugestões que devem ser consideradas por grandes/medias corporações antes e durante o processo de aquisição de uma strat-up: (i) A base de clientes da empresa comprada deve ser cuidadosamente considerada; (ii) um contato muito próximo entre os gestores das duas empresas é crucial antes da realização da aquisição; (iii) a contratação de uma empresa de consultoria em aquisições pode ser primordial durante o processo de integração; (iv) o empreendedor tem um papel de central importância para o future da nova empresa formada após a aquisição; (v) a forma como a integração entre as duas empresas ocorrerá após a compra deve ser cuidadosamente escolhida e (iv) a criação de uma corporate venture deve ser levada em consideração.

Assessment of value creation in private equity: the acquisition of Burger King by 3G Capital

This thesis elaborates the creation of value in private equity and in particular analyzes value creation in 3G Capital’s acquisition of Burger King. In this sense, a specific model is applied that composes value creation into several drivers, in order to answer the question of how value creation can be addressed in private equity investments. Although previous research by Achleitner et al. (2010) introduced a specific model that addresses value creation in private equity, the respective model was neither applied to an individual company, nor linked to indirect drivers that explain the dynamics and rationales for the creation of value. In turn this paper applies the quantitative model to an ongoing private equity investment and thereby provides different extensions to turn the model into a better forecasting model for ongoing investments, instead of only analyzing a deal that has already been divested from an ex post perspective. The chosen research approach is a case study about the Burger King buyout that first includes an extensive review about the current status of academic literature, second a quantitative calculation and qualitative interpretation of different direct value drivers, third a qualitative breakdown of indirect drivers, and lastly a recapitulating discussion about value creation and value drivers. Presenting a very successful private equity investment and elaborately demonstrating the dynamics and mechanisms that drive value creation in this case, provides important implications for other private equity firms as well as public firms in order to develop their proprietary approach towards value creation.

Flexible information acquisition and optimal Tobin tax in tractable dynamic global games

My dissertation focuses on dynamic aspects of coordination processes such as reversibility of early actions, option to delay decisions, and learning of the environment from the observation of other people’s actions. This study proposes the use of tractable dynamic global games where players privately and passively learn about their actions’ true payoffs and are able to adjust early investment decisions to the arrival of new information to investigate the consequences of the presence of liquidity shocks to the performance of a Tobin tax as a policy intended to foster coordination success (chapter 1), and the adequacy of the use of a Tobin tax in order to reduce an economy’s vulnerability to sudden stops (chapter 2). Then, it analyzes players’ incentive to acquire costly information in a sequential decision setting (chapter 3). In chapter 1, a continuum of foreign agents decide whether to enter or not in an investment project. A fraction λ of them are hit by liquidity restrictions in a second period and are forced to withdraw early investment or precluded from investing in the interim period, depending on the actions they chose in the first period. Players not affected by the liquidity shock are able to revise early decisions. Coordination success is increasing in the aggregate investment and decreasing in the aggregate volume of capital exit. Without liquidity shocks, aggregate investment is (in a pivotal contingency) invariant to frictions like a tax on short term capitals. In this case, a Tobin tax always increases success incidence. In the presence of liquidity shocks, this invariance result no longer holds in equilibrium. A Tobin tax becomes harmful to aggregate investment, which may reduces success incidence if the economy does not benefit enough from avoiding capital reversals. It is shown that the Tobin tax that maximizes the ex-ante probability of successfully coordinated investment is decreasing in the liquidity shock. Chapter 2 studies the effects of a Tobin tax in the same setting of the global game model proposed in chapter 1, with the exception that the liquidity shock is considered stochastic, i.e, there is also aggregate uncertainty about the extension of the liquidity restrictions. It identifies conditions under which, in the unique equilibrium of the model with low probability of liquidity shocks but large dry-ups, a Tobin tax is welfare improving, helping agents to coordinate on the good outcome. The model provides a rationale for a Tobin tax on economies that are prone to sudden stops. The optimal Tobin tax tends to be larger when capital reversals are more harmful and when the fraction of agents hit by liquidity shocks is smaller. Chapter 3 focuses on information acquisition in a sequential decision game with payoff complementar- ity and information externality. When information is cheap relatively to players’ incentive to coordinate actions, only the first player chooses to process information; the second player learns about the true payoff distribution from the observation of the first player’s decision and follows her action. Miscoordination requires that both players privately precess information, which tends to happen when it is expensive and the prior knowledge about the distribution of the payoffs has a large variance.

The status of soil phosphate fractions and the ability of fungi to dissolve hardly soluble phosphates

Brazilian soils predominantly consist of iron and aluminum oxides and have a low phosphorus content. The present study was carried out in order to assess the status of phosphate fractions in pasture, forest and agricultural soils and the ability of soil fungi to solubilize iron and aluminum phosphates. The abundance of P fractions in the soils studied occurred in the following order: Fe-P > reductant-soluble Fe-P > occluded Fe-P > occluded Al-P > Al-P > Ca-P. of the 481 fungi isolated, 33 showed the ability to solubilize the inorganic phosphates in culture. of these, 14 were considered to be high or very high solubilizers based on a solubilization capacity > 1000 mu g PO43- ml(-1). Isolate F-111 was the only one that dissolved all the insoluble phosphates used. Nine isolates solubilized both Al-P and Ca-P, and four other isolates only solubilized Ca-P. The highest number of isolates with high solubilization capacity were detected in pasture soil, followed by tropical rain forest and forest patch soils. Pasture soil presented both the largest contents of insoluble phosphates and the largest number of fungal isolates with phosphate-solubilizing ability. The range and size of P fractions influenced the number of fungi and their ability to solubilize hardly soluble phosphates. (c) 2004 Elsevier B.V. All rights reserved.

Bacterial diversity in soil in response to different plans, phosphate fertilizers and liming

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

Effectiveness of fused magnesium potassium phosphate for marandu grass

O alto custo atual do KCl e a grande dependência de sua importação para suprir a demanda nacional sugerem a necessidade de estudos que procurem avaliar a eficiência de outras fontes de K, principalmente aquelas baseadas em matéria-prima nacional. Nesse sentido, foi conduzido um experimento em casa de vegetação com amostras de um Latossolo Vermelho distrófico textura média, adotando-se o delineamento inteiramente casualizado em esquema fatorial 4 x 3 x 2, sendo quatro doses de K (0, 60, 120 e 180 mg kg-1 de K), três fontes [(cloreto de potássio (KCl)], termofosfato magnesiano potássico (FMPP) e a mistura de 70 % FMPP + 30 % KCl) e duas granulometrias (100 e 60 mesh), com três repetições. Verificou-se que a adubação potássica promoveu incrementos significativos na produção de matéria seca (parte aérea) e nos teores de K no solo e na planta, não havendo diferenças entre as fontes e suas granulometrias. Os níveis críticos de K no solo e na parte aérea das plantas foram de 1,53 mmol c dm-3 e 19,1 g kg-1, respectivamente.