Quantitative comparisons and models of time-averaging in bivalve and brachiopod shell accumulations

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

Human granulocyte colony stimulating factor (hG-CSF): cloning, overexpression, purification and characterization

Background: Biopharmaceutical drugs are mainly recombinant proteins produced by biotechnological tools. The patents of many biopharmaceuticals have expired, and biosimilars are thus currently being developed. Human granulocyte colony stimulating factor (hG-CSF) is a hematopoietic cytokine that acts on cells of the neutrophil lineage causing proliferation and differentiation of committed precursor cells and activation of mature neutrophils. Recombinant hG-CSF has been produced in genetically engineered Escherichia coli ( Filgrastim) and successfully used to treat cancer patients suffering from chemotherapy-induced neutropenia. Filgrastim is a 175 amino acid protein, containing an extra N-terminal methionine, which is needed for expression in E. coli. Here we describe a simple and low-cost process that is amenable to scaling-up for the production and purification of homogeneous and active recombinant hG-CSF expressed in E. coli cells.Results: Here we describe cloning of the human granulocyte colony-stimulating factor coding DNA sequence, protein expression in E. coli BL21(DE3) host cells in the absence of isopropyl-beta-D-thiogalactopyranoside ( IPTG) induction, efficient isolation and solubilization of inclusion bodies by a multi-step washing procedure, and a purification protocol using a single cationic exchange column. Characterization of homogeneous rhG-CSF by size exclusion and reverse phase chromatography showed similar yields to the standard. The immunoassay and N-terminal sequencing confirmed the identity of rhG-CSF. The biological activity assay, in vivo, showed an equivalent biological effect (109.4%) to the standard reference rhG-CSF. The homogeneous rhG-CSF protein yield was 3.2 mg of bioactive protein per liter of cell culture.Conclusion: The recombinant protein expression in the absence of IPTG induction is advantageous since cost is reduced, and the protein purification protocol using a single chromatographic step should reduce cost even further for large scale production. The physicochemical, immunological and biological analyses showed that this protocol can be useful to develop therapeutic bioproducts. In summary, the combination of different experimental strategies presented here allowed an efficient and cost-effective protocol for rhG-CSF production. These data may be of interest to biopharmaceutical companies interested in developing biosimilars and healthcare community.

Molecular models of NS3 protease variants of the Hepatitis C virus

Background: Hepatitis C virus (HCV) currently infects approximately three percent of the world population. In view of the lack of vaccines against HCV, there is an urgent need for an efficient treatment of the disease by an effective antiviral drug. Rational drug design has not been the primary way for discovering major therapeutics. Nevertheless, there are reports of success in the development of inhibitor using a structure-based approach. One of the possible targets for drug development against HCV is the NS3 protease variants. Based on the three-dimensional structure of these variants we expect to identify new NS3 protease inhibitors. In order to speed up the modeling process all NS3 protease variant models were generated in a Beowulf cluster. The potential of the structural bioinformatics for development of new antiviral drugs is discussed.Results: the atomic coordinates of crystallographic structure 1CU1 and 1DY9 were used as starting model for modeling of the NS3 protease variant structures. The NS3 protease variant structures are composed of six subdomains, which occur in sequence along the polypeptide chain. The protease domain exhibits the dual beta-barrel fold that is common among members of the chymotrypsin serine protease family. The helicase domain contains two structurally related beta-alpha-beta subdomains and a third subdomain of seven helices and three short beta strands. The latter domain is usually referred to as the helicase alpha-helical subdomain. The rmsd value of bond lengths and bond angles, the average G-factor and Verify 3D values are presented for NS3 protease variant structures.Conclusions: This project increases the certainty that homology modeling is an useful tool in structural biology and that it can be very valuable in annotating genome sequence information and contributing to structural and functional genomics from virus. The structural models will be used to guide future efforts in the structure-based drug design of a new generation of NS3 protease variants inhibitors. All models in the database are publicly accessible via our interactive website, providing us with large amount of structural models for use in protein-ligand docking analysis.

The water factor in the protein-folding problem

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

Effects of cement thickness and bonding on the failure loads of CAD/CAM ceramic crowns: Multi-physics FEA modeling and monotonic testing

Objective. To determine the influence of cement thickness and ceramic/cement bonding on stresses and failure of CAD/CAM crowns, using both multi-physics finite element analysis and monotonic testing.Methods. Axially symmetric FEA models were created for stress analysis of a stylized monolithic crown having resin cement thicknesses from 50 to 500 mu m under occlusal loading. Ceramic-cement interface was modeled as bonded or not-bonded (cement-dentin as bonded). Cement polymerization shrinkage was simulated as a thermal contraction. Loads necessary to reach stresses for radial cracking from the intaglio surface were calculated by FEA. Experimentally, feldspathic CAD/CAM crowns based on the FEA model were machined having different occlusal cementation spaces, etched and cemented to dentin analogs. Non-bonding of etched ceramic was achieved using a thin layer of poly(dimethylsiloxane). Crowns were loaded to failure at 5 N/s, with radial cracks detected acoustically.Results. Failure loads depended on the bonding condition and the cement thickness for both FEA and physical testing. Average fracture loads for bonded crowns were: 673.5 N at 50 mu m cement and 300.6 N at 500 mu m. FEA stresses due to polymerization shrinkage increased with the cement thickness overwhelming the protective effect of bonding, as was also seen experimentally. At 50 mu m cement thickness, bonded crowns withstood at least twice the load before failure than non-bonded crowns.Significance. Occlusal "fit" can have structural implications for CAD/CAM crowns; pre-cementation spaces around 50-100 mu m being recommended from this study. Bonding benefits were lost at thickness approaching 450-500 mu m due to polymerization shrinkage stresses. (C) 2012 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

Pion form factor in the light-front

The pion electromagnetic form factor is calculated with a light-front quark model. The plus and minus components of the electromagnetic current are used to calculate the electromagnetic form factor in the the Breit frame with two models for the q (q) over bar vertex. The light-front constituent quark model describes very well the hadronic wave functions for pseudo-scalar and vector particles. Symmetry problems arising in the light-front approcah are solved by the pole dislocation method. The results are compared with new experimental data and with other quark models.

Dynamically induced spontaneous symmetry breaking in 3-3-1 models

We show that in SU(3)(C) circle times SU(3)(L) circle times U(1)(N) (3-3-1) models embedded with a singlet scalar playing the role of the axion, after imposing scale invariance, the breaking of Peccei-Quinn symmetry occurs through the one-loop effective potential for the singlet field. We, then, analyze the structure of spontaneous symmetry breaking by studying the new scalar potential for the model, and verify that electroweak symmetry breaking is tightly connected to the 3-3-1 breaking by the strong constraints among their vacuum expectation values. This offers a valuable guide to write down the correct pattern of symmetry breaking for multi-scalar theories. We also obtained that the accompanying massive pseudo-scalar, instead of acquiring mass of order of Peccei-Quinn scale as we would expect, develops a mass at a much lower scale, a consequence solely of the breaking via Coleman-Weinberg mechanism. (c) 2005 Published by Elsevier B.V.

On high Q(2) behavior of the pion form factor for transitions gamma*gamma -> pi(0) and gamma*gamma* -> pi(0) within the nonlocal quark-pion model

The behavior of the transition pion form factor for processes gamma (*)gamma --> pi(0) and gamma (*)gamma (*) --> pi(0) at large values of space-like photon momenta is estimated within the nonlocal covariant quark-pion model. It is shown that, in general, the coefficient of the leading asymptotic term depends dynamically on the ratio of the constituent quark mass and the average virtuality of quarks in the vacuum and kinematically on the ratio of photon virtualities. The kinematic dependence of the transition form factor allows us to obtain the relation between the pion light-cone distribution amplitude and the quark-pion vertex function. The dynamic dependence indicates that the transition form factor gamma (*)gamma -->, pi(0) at high momentum transfers is very sensitive to the nonlocality size of nonperturbative fluctuations in the QCD vacuum. (C) 2000 Elsevier B.V. B.V. All rights reserved.

Flavor changing interaction models and strictly massless neutrinos

It is well known that experimental data, coming from solar and atmospheric neutrino detectors and also from experiments which look for neutrino oscillations. strongly suggest that neutrinos must have a mass different from zero. However at least the solar and/or the atmospheric neutrino data can be related to new flavor changing interactions beyond the standard model instead to the finite mass of neutrinos. This new physics may induce i) extra effects in neutrino-matter interactions, ii) CP violation in pion and lepton decays and, iii) muonium to antimuonium transition. We give two examples of models in which all those effects arise even with strictly massless neutrinos: the 331 model and multi-Higgs doublet extension of the standard model (mHDM) with flavor changing neutral currents in the charged lepton sector. It means that in this kind of models if neutrino masses were eventually needed, they will be independent of the parameters of the new interactions.

Degeneration of the Y chromosome in evolutionary aging models

The Y chromosomes are genetically degenerated and do not recombine with their matching partners X. Recombination of XX pairs is pointed out as the key factor for the Y chromosome degeneration. However, there is an additional evolutionary force driving sex-chromosomes evolution. Here we show this mechanism by means of two different evolutionary models, in which sex chromosomes with non-recombining XX and XY pairs of chromosomes is considered. Our results show three curious effects. First, we observed that even when both XX and XY pairs of chromosomes do not recombine, the Y chromosomes still degenerate. Second, the accumulation of mutations on Y chromosomes followed a completely different pattern then those accumulated on X chromosomes. and third, the models may differ with respect to sexual proportion. These findings suggest that a more primeval mechanism rules the evolution of Y chromosomes due exclusively to the sex-chromosomes asymmetry itself, i.e., the fact that Y chromosomes never experience female bodies. Over aeons, natural selection favored X chromosomes spontaneously, even if at the very beginning of evolution, both XX and XY pairs of chromosomes did not recombine.

Kaon and Pion Electromagnetic Form Factor Ratios in the Light-Front

We have applied the light-front formalism to calculate the electromagnetic form factors for the pion and the kaon from two models at low and high energies in order to explore the differences between such models. We have also compared the results for the ratio F(K)(Q(2))/F(pi)(Q(2)) with the experimental data, up to 10 [GeV/c](2) and we have observed that the theoretical results are in good concordance for low energies, but they are very different at higher energy scales.

Mini-implant and Nance button for initial retraction of maxillary canines: a prospective study in cast models

OBJETIVO: a ancoragem óssea é fundamental para o sucesso do tratamento de algumas más oclusões, pois permite a aplicação de forças contínuas, diminui o tempo de tratamento e independe da colaboração do paciente. MÉTODOS: o propósito desse trabalho foi comparar, por meio de modelos dentários, a perda de ancoragem após a retração inicial de caninos superiores entre dois grupos. O grupo A utilizou o mini-implante enquanto o grupo B utilizou o Botão de Nance. Para todos os pacientes foram realizados dois modelos (M1 e M2). Os primeiros modelos foram realizados ao início (M1), e os outros ao final da retração inicial de canino (M2). RESULTADOS: todas as medidas foram tabuladas e submetidas à análise estatística. Para verificar o erro sistemático intraexaminador foi utilizado o teste t pareado. Na determinação do erro casual utilizou-se o cálculo de erro proposto por Dahlberg. Para comparação entre as fases Início e Após, foi utilizado o teste t pareado. Para a comparação entre os grupos de mini-implante e Botão de Nance, foi utilizado o teste t de Student para medidas independentes. em todos os testes foi adotado nível de significância de 5% (p<0,05). CONCLUSÃO: ao se medir e comparar em modelos dentários a perda de ancoragem dos molares após a retração inicial de canino utilizando-se dois sistemas de ancoragem distintos (Mini-implante e Botão de Nance), pôde-se observar a inexistência de diferença estatisticamente significativa entre os dois grupos.

A multi-cell variable frequency interleaved ZCS boost rectifier digitally controlled by FPGA

This paper presents a multi-cell single-phase high power factor boost rectifier in interleave connection, operating in critical conduction mode, employing a soft-switching technique, and controlled by Field Programmable Gate Array (FPGA). The soft-switching technique is based on zero-current-switching (ZCS) cells, providing ZC (zero-current) turn-on and ZCZV (zero-current-zero-voltage) turn-off for the active switches, and ZV (zero-vohage) turn-on and ZC (zero-current) turn-off for the boost diodes. The disadvantages related to reverse recovery effects of boost diodes operated in continuous conduction mode (additional losses, and electromagnetic interference (EMI) problems) are minimized, due to the operation in critical conduction mode. In addition, due to the interleaving technique, the rectifier's features include the reduction in the input current ripple, the reduction in the output voltage ripple, the use of low stress devices, low volume for the EMI input filter, high input power factor (PF), and low total harmonic distortion (THD) in the input current, in compliance with the IEC61000-3-2 standards. The digital controller has been developed using a hardware description language (VHDL) and implemented using a XC2S200E-SpartanII-E/Xilinx FPGA device, performing a true critical conduction operation mode for all interleaved cells, and a closed-loop to provide the output voltage regulation, like as a preregulator rectifier. Experimental results are presented for a implemented prototype with two and with four interleaved cells, 400V nominal output voltage and 220V(rms) nominal input voltage, in order to verify the feasibility and performance of the proposed digital control through the use of a FPGA device.

A phase-shifting control for variable frequency multi-cells interleaved boost pre-regulator based on FPGA device

In this paper were investigated phase-shift control strategies applied to a four cells interleaved high input-power-factor pre-regulator boost rectifier, operating in critical conduction mode, using a non-dissipative commutation cells and frequency modulation. The digital control has been developed using a hardware description language (VHDL) and implemented using the XC2S200E-SpartanII-E/Xilinx FPGA, performing a true critical conduction operation mode for a generic number of interleaved cells. Experimental results are presented, in order to verify the feasibility and performance of the proposed digital control, through the use of a Xilinx FPGA device.

Analysis of the Stayability in Milk Buffaloes using Survival Models

In this study the trait Stayability (SA) was evaluated according to the year of cull after first calvin, i.e., SA 1 to 6 for 1 to 6 years from first calving in lactating females from bubaline milk herds spread in nine farms located in São Paulo state. Informations were used regarding 1027 lactating Murrah breed buffaloes. The statistical analyses were made using LIFEREG (SAS, 1999) procedure. The SA was evaluated using the fixed effects: farm production, birth year, calving season (Season 1- April to September and Season 2 October - March) and class of milk yield at 270 days. The age at first calving (AFC) was considered as a random effect. The mean observed for total milk yield was 1458.75Kg. Calving Season 2 encloses 65.6% of births. The means of cull age, in months, and the percentage of SA were, respectively: 10.69 e 69% (SA1), 19.30 e 63% (SA2), 26.4 e 54% (SA3), 33.15 e 42% (SA4), 38.53 e 36% (SA5) e 42.65 e 26% (SA6). It is verified that most of culls happens after the first lactation, among the sixth and eleventh month after first calving. It was observed that the factors: farm production, birth year and class of milk yield at 270 days affected significantly all SAs. Factors like calving season and the age at first calving (AFC) were only significant for SAL Being significant the factor AFC in level of 1% and factor time in 10%. For other SAs these factors were not statistically significant.