Structure and Mode of Action of Microplusin, a Copper II-chelating Antimicrobial Peptide from the Cattle Tick Rhipicephalus (Boophilus) microplus

Microplusin, a Rhipicephalus (Boophilus) microplus antimicrobial peptide (AMP) is the first fully characterized member of a new family of cysteine-rich AMPs with histidine-rich regions at the N and C termini. In the tick, microplusin belongs to the arsenal of innate defense molecules active against bacteria and fungi. Here we describe the NMR solution structure of microplusin and demonstrate that the protein binds copper II and iron II. Structured as a single alpha-helical globular domain, microplusin consists of five alpha-helices: alpha 1 (residues Gly-9 to Arg-21), alpha 2 (residues Glu-27 to Asn-40), alpha 3 (residues Arg-44 to Thr-54), alpha 4 (residues Leu-57 to Tyr-64), and alpha 5 (residues Asn-67 to Cys-80). The N and C termini are disordered. This structure is unlike any other AMP structures described to date. We also used NMR spectroscopy to map the copper binding region on microplusin. Finally, using the Gram-positive bacteria Micrococcus luteus as a model, we studied of mode of action of microplusin. Microplusin has a bacteriostatic effect and does not permeabilize the bacterial membrane. Because microplusin binds metals, we tested whether this was related to its antimicrobial activity. We found that the bacteriostatic effect of microplusin was fully reversed by supplementation of culture media with copper II but not iron II. We also demonstrated that microplusin affects M. luteus respiration, a copper-dependent process. Thus, we conclude that the antibacterial effect of microplusin is due to its ability to bind and sequester copper II.

Combining structure and dynamics: non-denaturing high-pressure effect on lysozyme in solution

Small-angle X-ray scattering (SAXS) and elastic and quasi-elastic neutron scattering techniques were used to investigate the high-pressure-induced changes on interactions, the low-resolution structure and the dynamics of lysozyme in solution. SAXS data, analysed using a global-fit procedure based on a new approach for hydrated protein form factor description, indicate that lysozyme completely maintains its globular structure up to 1500 bar, but significant modi. cations in the protein-protein interaction potential occur at approximately 600-1000 bar. Moreover, the mass density of the protein hydration water shows a clear discontinuity within this pressure range. Neutron scattering experiments indicate that the global and the local lysozyme dynamics change at a similar threshold pressure. A clear evolution of the internal protein dynamics from diffusing to more localized motions has also been probed. Protein structure and dynamics results have then been discussed in the context of protein-water interface and hydration water dynamics. According to SAXS results, the new configuration of water in the first hydration layer induced by pressure is suggested to be at the origin of the observed local mobility changes.

Cearania heterophylla gen. nov et sp nov., a fossil gymnosperm with affinities to the Gnetales from the Early Cretaceous of northern Gondwana

Vegetative and fertile shoots of a shrub-like seed plant from the late Aptian Crato Formation of Brazil are described as Cearania heterophylla Kunzmann, Mohr and Bernardes-de-Oliveira, gen. nov. et sp. nov. Anatomical details of the axes, epidermal features and separate ovulate and pollen producing organs indicate the gymnospermous nature of this plant. The vascular tissue of the axes includes tracheids with bordered pits and fiber tracheids. Vegetative shoots comprising at least three branching orders bear opposite-decussately arranged ovate to lanceolate, dorsiventrally flattened, parallelodromous, rather thick leaves that vary tremendously in size. The amphistomatic leaves bear (brachy-)paracytic stomatal complexes arranged in simple longitudinal files. The ovulate structure is interpreted as a terminally attached single globular ovule/seed surrounded by at least five to six lanceolate bracts. A terminally attached pollen-cone like structure grows on a lateral leafy shoot. The unusual character combination may indicate that the fossils belong to a hitherto unknown group with affinities to ephedroid Gnetales. Sterile shoots formerly often described as Podozamites, Nageiopsis or Lilites that are at least partly congeneric with C. heterophylla Kunzmann, Mohr and Bernardes-de-Oliveira, gen. nov. et sp. nov. had a wide geographic distribution during the Early Cretaceous. (C) 2009 Elsevier B.V. All rights reserved.

Structure, processing and midgut secretion of putative peritrophic membrane ancillary protein (PMAP) from Tenebrio molitor larvae

A cDNA coding for a Tenebrio molitor midgut protein named peritrophic membrane ancillary protein (PMAP) was cloned and sequenced. The complete cDNA codes for a protein of 595 amino acids with six insect-allergen-related-repeats that may be grouped in A (predicted globular)- and B (predicted nonglobular)-types forming an ABABAB structure. The PMAP-cDNA was expressed in Pichia pastoris and the recombinant protein (64 kDa) was purified to homogeneity and used to raise antibodies in rabbits. The specific antibody detected PMAP peptides (22 kDa) in the anterior and middle midgut tissue, luminal contents, peritrophic membrane and feces. These peptides derive from PMAP, as supported by mass spectrometry, and resemble those formed by the in vitro action of trypsin on recombinant PMAP. Both in vitro and in vivo PMAP processing seem to occur by attack of trypsin to susceptible bonds in the coils predicted to link AB pairs, thus releasing the putative functional AB structures. The AB-domain structure of PMAP is found in homologous proteins from several insect orders, except lepidopterans that have the apparently derived protein known as nitrile-specifier protein. Immunocytolocalization shows that PMAP is secreted by exocytosis and becomes entrapped in the glycocalyx, before being released into midgut contents. Circumstantial evidence suggests that PMAP-like proteins have a role in peritrophic membrane type 2 formation. (C) 2007 Elsevier Ltd. All rights reserved.

Revision of tertiary species of Anaskopora Wass (Bryozoa: Cribrimorpha)

The subgenus Anaskopora Wass, 1975 is raised to generic rank, separated from the genus Corbulipora, and redefined. The type species, Cribrilina elevata MacGillivray, 1895, is a Tertiary fossil from Victoria with small globular colonies formed principally by a special kind of interzooidal frontal budding. Other Tertiary fossil species with a similar colony structure, here assigned to Anaskopora, are Cribrilma cornuta MacGIllIvray, 1895 and Lepralia rotundata MacGillivray, 1895. Two further new Tertiary species, A. simplex and A. mesa, from Victoria and South Australia have small encrusting colonies. A key to species is given.

Differential regulation of Adiponectin receptor gene expression by Adiponectin and Leptin in Myotubes derived from obese and diabetic individuals

Objective: This study aimed to investigate the regulation of adiponectin receptors 1 (AdipoR1) and 2 (AdipoR2) gene expression in primary skeletal muscle myotubes, derived from human donors, after exposure to globular adiponectin (gAd) and leptin. Research Methods and Procedures: Four distinct primary cell culture groups were established [ Lean, Obese, Diabetic, Weight Loss (Wt Loss); n = 7 in each] from rectus abdominus muscle biopsies obtained from surgical patients. Differentiated myotube cultures were exposed to gAd (0.1 mug/mL) or leptin (2.5 mug/mL) for 6 hours. AdipoR1 and AdipoR2 gene expression was measured by real-time polymerase chain reaction analysis. Results: AdipoR1 mRNA expression in skeletal muscle myotubes derived from Lean subjects (p < 0.05) was stimulated 1.8-fold and 2.5-fold with gAd and leptin, respectively. No increase in AdipoR1 gene expression was measured in myotubes derived from Obese, Diabetic, or Wt Loss subjects. AdipoR2 mRNA expression was unaltered after gAd and leptin exposure in all myotube groups. Discussion: Adiponectin and leptin are rapid and potent stimulators of AdipoR1 in myotubes derived from lean healthy individuals. This effect was abolished in myotubes derived from obese, obese diabetic subjects, and obese-prone individuals who had lost significant weight after bariatric surgery. The incapacity of skeletal muscle of obese and diabetic individuals to respond to exogenous adiponectin and leptin may be further suppressed as a result of impaired regulation of the AdipoR1 gene.

Adiponectin decreases pyruvate dehydrogenase kinase 4 gene expression in obese- and diabetic derived

Aim: To investigate the effects of globular adiponectin (gAd) on gene expression and whether these effects are mediated through 3',5'-cyclic monophosphate-activated protein kinase in skeletal muscle myotubes obtained from lean, obese and obese diabetic individuals.

Methods: Rectus abdominus muscle biopsies were obtained from surgical patients to establish primary skeletal muscle cell cultures. Three distinct primary cell culture groups were established (lean, obese and obese diabetic; n = 7 in each group). Once differentiated, these cultures were then exposed to gAd or 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside (AICAR) for 6 h.

Results: Stimulation with gAd decreased pyruvate dehydrogenase kinase 4 (PDK4) gene expression in the obese and diabetic samples (p ≤ 0.05) and increased cytochrome c oxidase (COX) subunit 4 (COXIV) gene expression in the myotubes derived from lean individuals only (p < 0.05). AICAR treatment also decreased PDK4 gene expression in the obese- and diabetic-derived myotubes (p ≤ 0.05) and increased the gene expression of the mitochondrial gene, COXIII, in the lean-derived samples only (p < 0.05).

Conclusions: This study demonstrated distinct disparity between myotubes derived from lean compared with obese and obese diabetic individuals following gAd and AICAR treatment. Further understanding of the regulation of PDK4 in obese and diabetic skeletal muscle and its interaction with adiponectin signalling is required as this appears to be an important early molecular event in these disease states that may improve blood glucose control and metabolic flux.

Secondary emission of nanocrystalline zinc oxide

The Raman and photoluminescence (PL) spectra of nanocrystalline zinc oxide produced by mechanochemical synthesis were measured using a pulsed nitrogen laser (337.1 nm) and xenon lamp (360 nm) as excitation sources in PL measurements and a cw Nd:YAG laser in Raman measurements. PL was observed in the range 400–800 nm. The Raman spectrum of nanocrystalline (90 nm) ZnO was compared to that of coarsegrained ZnO. The Raman bands of nanocrystalline zinc oxide were found to be shifted to lower frequencies and broadened. Laser radiation was shown to cause local heating of zinc oxide up to 1000 K, resulting in photoinduced formation of zinc nanoclusters. Mixtures of zinc oxide and sodium chloride powders are heated to substantially lower temperatures. Under nitrogen laser excitation, the green PL band (535 nm), characteristic of bulk ZnO, is shifted to longer wavelengths by 85 nm. The results are interpreted in terms of light confinement in zinc oxide microclusters consisting of large number of nanocrystallites. The photoinduced processes in question may be a viable approach to producing metal-insulator structures in globular photonic crystals, opals, filled with zinc oxide.

Identifying foldable regions in protein sequence from the hydrophobic signal

Structural genomics initiatives aim to elucidate representative 3D structures for the majority of protein families over the next decade, but many obstacles must be overcome. The correct design of constructs is extremely important since many proteins will be too large or contain unstructured regions and will not be amenable to crystallization. It is therefore essential to identify regions in protein sequences that are likely to be suitable for structural study. Scooby-Domain is a fast and simple method to identify globular domains in protein sequences. Domains are compact units of protein structure and their correct delineation will aid structural elucidation through a divide-and-conquer approach. Scooby-Domain predictions are based on the observed lengths and hydrophobicities of domains from proteins with known tertiary structure. The prediction method employs an A*-search to identify sequence regions that form a globular structure and those that are unstructured. On a test set of 173 proteins with consensus CATH and SCOP domain definitions, Scooby-Domain has a sensitivity of 50% and an accuracy of 29%, which is better than current state-of-the-art methods. The method does not rely on homology searches and, therefore, can identify previously unknown domains.

Lubricin: a versatile, biological anti-adhesive with properties comparable to polyethylene glycol

Lubricin is a glycoprotein found in articular joints which has been recognized as being an important biological boundary lubricant molecule. Besides providing lubrication, we demonstrate, using a quartz crystal microbalance, that lubricin also exhibits anti-adhesive properties and is highly effective at preventing the non-specific adsorption of representative globular proteins and constituents of blood plasma. This impressive anti-adhesive property, combined with lubricin's ability to readily self-assemble to form dense, highly stable telechelic polymer brush layers on virtually any substrates, and its innate biocompatibility, makes it an attractive candidate for anti-adhesive and anti-fouling coatings. We show that coatings of lubricin protein are as effective as, or better than, self-assembled monolayers of polyethylene glycol over a wide range of pH and that this provides a simple, versatile, highly stable, and highly effective method of controlling unwanted adhesion to surfaces.

Gestão das relações intergovernamentais: fatores condicionantes da propensão a cooperar por parte dos governos locais

Dissertação apresentada ao Programa de Mestrado em Administração da Universidade Municipal de São Caetano do Sul - USCS

Os canais de distribuição como parte do composto de marketing adotado na comercialização de produtos evangélicos: fatores de sucesso da Rua Conde de Sarzedas

SOUZA, Roberto Vidal de. Os canais de distribuição como parte do composto de marketing adotado na comercialização de produtos evangélicos: fatores de sucesso da Rua Conde de Sarzedas. 2013. 134f. Dissertação (Mestrado em Administração)-Universidade Municipal de São Caetano do Sul, São Caetano do Sul, 2013.

Competitividade de clusters comerciais: estudo sobre uma aglomeração de pequenas lojas de veículos no município de São Paulo

O presente trabalho teve por objetivo identificar se a aglomeração de lojas de veículos existente na Avenida Marechal Tito / Avenida São Miguel (zona leste do município de São Paulo) forma um cluster comercial varejista, a partir dos fundamentos da performance competitiva de clusters descritos por Zaccarelli et al. (2008). Este é um estudo descritivo de natureza quantitativa, executado a partir de amostragem não-probabilística fundamentada no critério de acessibilidade dos pesquisadores junto aos proprietários e gestores de 128 lojas de automóveis, das quais 85,94% são microempresas, 12,50% são de pequeno porte e 1,56% são de médio porte. Constatou-se que a localidade ainda se encontra em um estágio inicial de cluster comercial varejista, pois apesar de haver forte concorrência entre os lojistas naquela concentração geográfica, com ausência de posições privilegiadas e substituição seletiva dos participantes não competitivos, a cooperação entre eles é incipiente, bem como não há nenhuma iniciativa voltada para o aumento da competitividade do aglomerado como um todo.

Clusters como estratégia de desenvolvimento de pequenas e médias empresas: um estudo exploratório do caso moveleiro do município de Colatina - Es

Esse trabalho tem como objetivo mostrar que a formação de clusters de pequenas e médias empresas na área de móveis pode representar uma futura alternativa econômica para Colatina. o foco principal desse estudo visa, analisar o aglomerado de empresas moveleiras do município de Colatina, localizado no centro-oeste do Espírito Santo. Pretende-se identificar como estas empresas estão organizadas, quais barreiras são enfrentadas por elas ao buscar formas de garantir competitividade, e quais atividades poderiam ser alvos de estímulos por parte das instituições de fomento com vistas a transformar este pólo moveleiro em um cluster dinâmico e iniciar um processo de desenvolvimento local. Observamos que Colatina, constitui um caso interessante de estudo por se tratar de um aglomerado de PMEs que, mesmo tendo que enfrentar alguns obstáculos, está conseguindo dinamizar o sistema produtivo local e atrair investimentos.

Arranjo produtivo local: o caso de São João Del Rei-MG

O objetivo dessa dissertação foi investigar o processo de formação do Arranjo Produtivo Local (APL) de estanho em São João del-Rei - A7 Estanho, particularizando suas contribuições para o desenvolvimento local do município. O trabalho foi conduzido mediante uma discussão teórica sobre a situação das micro e pequenas empresas no Brasil, sobre a importância econômica e social dos arranjos produtivos locais, seus impactos no desenvolvimento regional e/ou local, principalmente de municípios sem infra-estrutura para atrair grandes empresas, e sobre políticas públicas que estão em andamento e que se voltam para o fomento desse tipo de aglomerado. Foram realizadas cerca de quatorze entrevistas no município de São João del-Rei: seis integrantes da Associação, uma funcionária, dois representantes das principais instituições parceiras, um representante do governo local e quatro membros da sociedade civil, que possuíam informações relevantes sobre a A7. O método empregado no tratamento dos dados foi a análise de conteúdo. Como resultados, no que diz respeito à análise dos programas que deram origem e/ou sustentação ao aglomerado, verificou-se que dois programas foram mais contundentes em suas contribuições à Associação: o Programa Empreender, da Federaminas, que permitiu a união dos empresários em torno de um objetivo comum, e o Projeto Via Design, do Sebrae, que possibilitou uma série de vantagens ao grupo, solidificando-o no mercado. No que se refere à participação dos diversos atores (membros da A7, poder público, instituições parceiras, sociedade civil) na construção do APL, pode-se perceber que o nível de atuação e comprometimento variava de acordo com as motivações pessoais, cargos ou funções formais exercidas e grau de relacionamento com a Associação. As contribuições ao desenvolvimento local do município, geradas pelo arranjo produtivo, foram enumeradas como geração de emprego e renda, ainda de forma incipiente, mas com possibilidades de crescimento futuro, e divulgação do turismo e cultura sãojoanense.