The development of chromatography for the characterization of protein interactions: a personal perspective

This article reviews the progress of a personal endeavour to develop chromatography as a quantitative procedure for the determination of reaction stoichiometries and equilibrium constants governing protein interactions. As well as affording insight into an aspect of chromatography with which many protein chemists are unfamiliar, it shows the way in which minor adaptations of conventional chromatographic practices have rendered the technique one of the most powerful methods available for the characterization of interactions. That pathway towards quantification is followed from the introduction of frontal gel filtration for the study of protein self-association to the characterization of ligand binding by the biosensor variant of quantitative affinity chromatography.

Model-based analysis of anaerobic acetate uptake by a mixed culture of polyphosphate-accumulating and glycogen-accumulating organisms

An increasing number of studies shows that the glycogen-accumulating organisms (GAOs) can survive and may indeed proliferate under the alternating anaerobic/aerobic conditions found in EBPR systems, thus forming a strong competitor of the polyphosphate-accumulating organisms (PAOs). Understanding their behaviors in a mixed PAO and GAO culture under various operational conditions is essential for developing operating strategies that disadvantage the growth of this group of unwanted organisms. A model-based data analysis method is developed in this paper for the study of the anaerobic PAO and GAO activities in a mixed PAO and GAO culture. The method primarily makes use of the hydrogen ion production rate and the carbon dioxide transfer rate resulting from the acetate uptake processes by PAOs and GAOs, measured with a recently developed titration and off-gas analysis (TOGA) sensor. The method is demonstrated using the data from a laboratory-scale sequencing batch reactor (SBR) operated under alternating anaerobic and aerobic conditions. The data analysis using the proposed method strongly indicates a coexistence of PAOs and GAOs in the system, which was independently confirmed by fluorescent in situ hybridization (FISH) measurement. The model-based analysis also allowed the identification of the respective acetate uptake rates by PAOs and GAOs, along with a number of kinetic and stoichiometric parameters involved in the PAO and GAO models. The excellent fit between the model predictions and the experimental data not involved in parameter identification shows that the parameter values found are reliable and accurate. It also demonstrates that the current anaerobic PAO and GAO models are able to accurately characterize the PAO/GAO mixed culture obtained in this study. This is of major importance as no pure culture of either PAOs or GAOs has been reported to date, and hence the current PAO and GAO models were developed for the interpretation of experimental results of mixed cultures. The proposed method is readily applicable for detailed investigations of the competition between PAOs and GAOs in enriched cultures. However, the fermentation of organic substrates carried out by ordinary heterotrophs needs to be accounted for when the method is applied to the study of PAO and GAO competition in full-scale sludges. (C) 2003 Wiley Periodicals, Inc.

Synthesis and characterization of delta-atracotoxin-ar1a, the lethal neurotoxin from venom of the Sydney funnel-web spider (Atrax robustus)

delta-Atracotoxin-Ar1a (delta-ACTX-Ar1a) is the major polypeptide neurotoxin isolated from the venom of the male Sydney funnel-web spider, Atrax robustus. This neurotoxin targets both insect and mammalian voltage-gated sodium channels, where it competes with scorpion alpha-toxins for neurotoxin receptor site-3 to slow sodium-channel inactivation. Progress in characterizing the structure and mechanism of action of this toxin has been hampered by the limited supply of pure toxin from natural sources. In this paper, we describe the first successful chemical synthesis and oxidative refolding of the four-disulfide bond containing delta-ACTX-Ar1a. This synthesis involved solid-phase Boc chemistry using double coupling, followed by oxidative folding of purified peptide using a buffer of 2 M GdnHCl and glutathione/glutathiol in a 1:1 mixture of 2-propanol (pH 8.5). Successful oxidation and refolding was confirmed using both chemical and pharmacological characterization. Ion spray mass spectrometry was employed to confirm the molecular weight. H-1 NMR analysis showed identical chemical shifts for native and synthetic toxins, indicating that the synthetic toxin adopts the native fold. Pharmacological studies employing whole-cell patch clamp recordings from rat dorsal root ganglion neurons confirmed that synthetic delta-ACTX-Ar1a produced a slowing of the sodium current inactivation and hyperpolarizing shifts in the voltage-dependence of activation and inactivation similar to native toxin. Under current clamp conditions, we show for the first time that delta-ACTX-Ar1a produces spontaneous repetitive plateau potentials underlying the clinical symptoms seen during envenomation. This successful oxidative refolding of synthetic delta-ACTX-Ar1a paves the way for future structure-activity studies to determine the toxin pharmacophore.

O caráter político-pedagógico dos movimentos populares de bairro da Grande São Pedro: avanços e recuos sob o imperativo da ordem capitalista

Este trabalho tem como objeto de estudo as ações organizativas dos movimentos populares de bairro, enquanto práticas político-pedagógicas. Tomamos como foco, as associações de moradores da região da Grande São Pedro. Apesar de ter ficado conhecida na década de 1980 pela situação de miséria, os moradores dessa região, historicamente perpassada pelo fenômeno da segregação socioespacial, encontraram nas adversidades motivação para a organização do movimento popular como forma de construir sua história e afirmar sua cidadania. Totalmente transformada no que tange à sua urbanização, a região ainda hoje apresenta um cenário adverso para os moradores. Nesse sentido, a importância da organização dos moradores na atualidade é fundamental para lutar contra as situações de opressão que se impõem sobre os mesmos. Entretanto, o quadro que os atuais movimentos populares de bairro desta região apresenta é de reprodução de um modelo político-cultural marcado pelo individualismo, com traços peculiares da formação social brasileira - caracterizada por práticas de centralização, dependência e clientelismo político. Deve-se registrar que apesar de também trazerem traços de lutas e resistência, a forma como atualmente acontecem, com ações em níveis imediatos, esvaziadas de conteúdo político mais amplo, sem articulação com movimentos mais abrangentes, acabam se expressando em um “associativismo individualizado”. Desta feita, reiteramos neste trabalho a necessidade de a classe trabalhadora brasileira retomar, reconstruir espaços que contribuam com a formação de lideranças, com a qualificação das bases desses movimentos, para que seus protagonistas possam consolidar uma prática político-pedagógica fundamentada na educação popular e com isso venham adensar as lutas da classe trabalhadora e somar ao projeto de construção da contra-hegemonia, visando à transformação social. Buscar-se-ia neste sentido, a consolidação de um projeto social mais igualitário, justo e verdadeiramente democrático, no qual seja possível a emancipação humana.

Estudo comparativo do pH e do volume salivar em indivíduos com laringofaringite crônica por doença do refluxo gastroesofágica antes e após o tratamento

INTRODUÇÃO: A Doença do Refluxo Gastroesofágico (DRGE) é a doença digestiva mais prevalente da atualidade e, recentemente, tem sido implicada em uma gama de alterações do seguimento laringofaríngeo (RLF). No entanto, pouco se sabe dos mecanismos fisiopatológicos destas manifestações supraesofágicas da DRGE. Os achados clínicos contraditórios e recentes pesquisas sugerem haver deficiências na capacidade de defesa deste seguimento. Uma das principais responsáveis pela homeostase da mucosa oral e do trato digestivo é a saliva com seu conteúdo orgânico e inorgânico. Tanto alterações do pH quanto do volume salivar já foram correlacionados com os sintomas e sinais sugestivos da DRGE e RLF. Estudo recente de nossa autoria demonstra diminuição estatisticamente significante do pH salivar de indivíduos com RLF quando comparado a controles sem a doença. Outro estudo constatou correlação entre a redução do volume X pH da saliva em indivíduos com DRGE, estando esta redução diretamente relacionada aos níveis de pH esofágico constatados durante pH-metria esofágica de 24 horas. OBJETIVOS: Avaliar como se comportam o pH e volume da saliva em um mesmo indivíduo com DRGE e RLF antes e após o tratamento clínico. MATERIAL E MÉTODO: Vinte e três pacientes com RLF tiveram o pH e volume da saliva total testados antes e após receberem tratamento com droga bloqueadora de bomba de prótons durante 12 semanas. RESULTADOS: Houve uma diferença estatisticamente significante (p<0,001) entre o pH da saliva antes e após o tratamento, estando este maior após o controle clínico da doença. O volume de saliva no paciente tratado foi significativamente maior do que no paciente pré-tratamento (p=0.009). DISCUSSÃO: Os achados sugerem que o pH salivar é influenciado pela presença de refluxo gastroduodenal à região laringofaríngea. Caso estudos futuros com populações maiores realmente comprovem esta correlação, poderemos cogitar a possibilidade de usar a mensuração do pH salivar, que é feita de forma rápida e não invasiva, como um meio de diagnosticar e avaliar o comportamento e controle do Refluxo Laringofaríngeo.

Sources of Brazil's Counter-Hegemony

There have been two key initiatives taken in the last two decades in Brazil to create a counter-hegemonic project for the country. One initiative resulted from Brazil's business community and high-level State bureaucracy and aimed at forming a regional economic and political bloc that would guarantee and enlarge a relative independence from the hegemonic powers. The other resulted from the emergence of the new unionist movement in São Paulo and from the formation of Partido dos Trabalhadores and aimed at promoting radical democratization and reducing social exclusion. Both initiatives have created policies and changes that have converged to enhance Brazil's counter-hegemonic position as a regional and emerging power.

Sugarcane productivity correlated with physical-chemical attributes to create soil management zone

The socioeconomic importance of sugar cane in Brazil is unquestionable because it is the raw material for the production of ethanol and sugar. The accurate spatial intervention in the management of the crop, resulting zones of soil management, increases productivity as well as its agricultural yields. The spatial and Person's correlations between sugarcane attributes and physico-chemical attributes of a Typic Tropustalf were studied in the growing season of 2009, in Suzanápolis, State of São Paulo, Brazil (20°28'10'' S lat.; 50°49'20'' W long.), in order to obtain the one that best correlates with agricultural productivity. Thus, the geostatistical grid with 120 sampling points was installed to soil and data collection in a plot of 14.6 ha with second crop sugarcane. Due to their substantial and excellent linear and spatial correlations with the productivity of the sugarcane, the population of plants and the organic matter content of the soil, by evidencing substantial correlations, linear and spatial, with the productivity of sugarcane, were indicators of management zones strongly attached to such productivity.

Determinantes da adopção e desenvolvimento do Sistema de Contabilidade de Custos nos municípios Portugueses

The present study envisages to analyse and perceive change within the Portuguese local governments’ accounting information system. This change has occurred in recent years due to the development of the Cost Accounting System (CAS). Are there divergences between those that adopted and those that did not the CAS? As of today, to what extent is the CAS implemented? What factors do determine the level of development of the CAS amongst Portuguese local governments? This study has analysed several factors like the current legal framework, the local governments’ size, the financial independence and the level of political competitiveness. The empirical analysis focus upon the results as obtained by means of questionnaire which was applied to all the Portuguese Local Government (308). Although one may consider that its development is rather shallow, evidence unfolds that about 48% of the respondents have already adopted the CAS. It highlights that the pressure exerted by the current legal framework, namely the POCAL (Plano Oficial de Contabilidade das Autarquias Locais – Local Government Accounting Official Plan), the NLFL (Nova Lei das Finanças Locais – Local Finances New Law) and the RGTAL (Regime Geral de Taxas das Autarquias Locais – Local Government General Regime of Taxes), justifies that most of the local governments has adopted the new system of control and accounting information in recent years.The importance of information stemming from the CAS, thus bettering and refining the decision making process, is consistent with the positive evolution of this system, in the short-run. The institutions’ size as well as the level of financial independence and political competitiveness may also be pondered as most significant variables within this empirical analysis.

STRATEGIES FOR SUPERIOR PERFORMANCE IN RECESSIONS: PRO OR COUNTER-CYCLICAL?

Recessions are recurring events in which most firms suffer severe impacts while others are less affected or may even prosper. Strategic management has made little progress in understanding such performance differences. In a scenario of decreased demand, intensified competition, and higher uncertainty, most firms try to survive by pro-cyclically cutting costs and investments. But firms could take advantage of undervalued resources in the market to counter-cyclically invest in new business opportunities to overtake competitors. We survey Brazilian firms in various industries about the 2008-2009 recession and analyze data using PLS-SEM. We find that while most firms pro-cyclically reduce costs and investments in recessions, a counter-cyclical strategy of investing in opportunities created by changes in the market enables superior performance. Most successful are firms with a propensity to recognize opportunities, an entrepreneurial orientation to invest, and the flexibility to efficiently implement investments.

Current issues in Social Work Theory

São aqui analisados os discursos sobre as tendências gerais na teoria de Serviço Social. Estes discursos sobre o papel do Serviço Social são tratados no contexto da sociedade, bem como do cliente -assistente sócial -agência. A teoria da prática representa um discurso entre as perspectivas de Serviço Social nos seus aspectos transformacionais, terapêuticos e de ordem social. São também abordadas as suas várias combinações e interacções em todo o tipo de prática e organização de Serviço Social. O discurso sobre a eficácia baseia¬se em estudos comprovados da construção social, potenciação (empowerment), e realismo crítico ainda polémico. A prática reflectiva e crítica constitui os modelos actuais que interligam a teoria e a prática.

Current trends in the global tourism industry: evidence from the United States

Tourism is one of the largest U.S. industries, serving millions of international and domestic tourists yearly. Tourists visit the U.S. to see natural wonders, cities, historic landmarks, and entertainment venues. Americans seek similar attractions as well as recreation and vacation areas. Tourism competes in the global market, so it is important to understand current trends in the U.S. travel industry. Therefore, this article offers insight into important trends and suggests strategies for policy makers involved in the travel and tourism industry.

Vector competence experiments with Rocio virus and three mosquito species from the epidemic zone in Brazil

First-generation progeny of field-collected Psorophora ferox, Aedes scapularis, and Aedes serratus from the Rocio encephalitis epidemic zone in S.Paulo State, Brazil, were tested for vector competency in the laboratory. Psorophora ferox and Ae. scapularis are susceptible to per os infection with Rocio virus and can transmit the virus by bite following a suitable incubation period. Oral ID50S for the two species (10(4.1) and 10(4.3) Vero cell plaque forming units, respectively) did not differ significantly. Infection rates in Ae. serratus never exceeded 36%, and, consequently, an ID50 could not be calculated for this species. It is unlikely that Ae. serratus is an epidemiologically important vector of Rocio virus. The utility of an in vitro feeding technique for demonstrating virus transmission by infected mosquitoes and difficulties encountered in working with uncolonized progeny of field-collected mosquitoes are discussed.

Impact of flavor-changing neutral current top quark interactions on BR(t -> bW)

We study the effect that flavor-changing neutral current interactions of the top quark will have on the branching ratio of charged decays of the top quark. We have performed an integrated analysis using Tevatron and B-factories data and with just the further assumption that the Cabibbo-Kobayashi-Maskawa matrix is unitary, we can obtain very restrictive bounds on the strong and electroweak flavor-changing neutral current branching ratios Br(t -> qX)< 4.0x10(-4), where X is any vector boson and a sum in q=u, c is implied.