Relation between sequence and structure of HIV-1 protease inhibitor complexes: a model system for the analysis of protein flexibility.

The flexibility of different regions of HIV-1 protease was examined by using a database consisting of 73 X-ray structures that differ in terms of sequence, ligands or both. The root-mean-square differences of the backbone for the set of structures were shown to have the same variation with residue number as those obtained from molecular dynamics simulations, normal mode analyses and X-ray B-factors. This supports the idea that observed structural changes provide a measure of the inherent flexibility of the protein, although specific interactions between the protease and the ligand play a secondary role. The results suggest that the potential energy surface of the HIV-1 protease is characterized by many local minima with small energetic differences, some of which are sampled by the different X-ray structures of the HIV-1 protease complexes. Interdomain correlated motions were calculated from the structural fluctuations and the results were also in agreement with molecular dynamics simulations and normal mode analyses. Implications of the results for the drug-resistance engendered by mutations are discussed briefly.

Value added logistical support service. Part 1 : Trends and new concept model

The aim of `Valssi' study was to find out the service requirements in `business-to-business' (B2B) markets and to present a new logistics service concept, where the traditional logistics service is expanded with significant manufacturing and value addedfacilities. The traditional third-party logistics service providers are not necessarily able to offer services, which cover widely the needs of potential customers. The study has been outlined to spare part markets in metal industry. In the second phase of the Valssi-project the aim is to examine the economical conditions and potentiality of the new logistics business concept. This research report (Part 1) concentrates on examining current trends in global and domestic logistics markets. Based on detailed survey among the participating companies, the study presents basis for a new logistics business concept model. The developed concept consists of 12 different service modules, which are split into deeper details of processes. The integration of worldwide supplier and service provider network together with customer companies systems is a challenge. The report focuses on evaluating the requirements for the new business concept from the customer-companies point of view. The study paints an overall picture of distribution and service provider network including an abstract about the software and system integration possibilities. As a result of the survey, it can be concluded that thereis need for the new business concept among the participating companies, and a modular service concept meets the requirements of them, because the new sophisticated concept considers the specialities involved in spare part logistics in metal industry.

Neuropathic Pain Phenotype Does Not Involve the NLRP3 Inflammasome and Its End Product Interleukin-1β in the Mice Spared Nerve Injury Model.

The NACHT, LRR and PYD domains-containing protein 3 (NLRP3) inflammasome is one of the main sources of interleukin-1β (IL-1β) and is involved in several inflammatory-related pathologies. To date, its relationship with pain has not been studied in depth. The aim of our study was to elucidate the role of NLRP3 inflammasome and IL-1β production on neuropathic pain. Results showed that basal pain sensitivity is unaltered in NLRP3-/- mice as well as responses to formalin test. Spared nerve injury (SNI) surgery induced the development of mechanical allodynia and thermal hyperalgesia in a similar way in both genotypes and did not modify mRNA levels of the NLRP3 inflammasome components in the spinal cord. Intrathecal lipopolysaccharide (LPS) injection increases apoptosis-associated speck like protein (ASC), caspase-1 and IL-1β expression in both wildtype and NLRP3-/- mice. Those data suggest that NLRP3 is not involved in neuropathic pain and also that other sources of IL-1β are implicated in neuroinflammatory responses induced by LPS.

A new alternative method for testing skin irritation using a fresh human skin model

La dermatite irritative est décrite comme une réaction réversible, non immunologique caractérisée par des lésions d'aspect très variable, allant de la simple rougeur jusqu'à la formation de bulles voire d'une nécrose, accompagnée de prurit ou d'une sensation de brûlure suite à I' application d'une substance chimique. Le teste de prédiction d'irritation cutanée est traditionnellement depuis les années 1940 le Test de Draize. Ce test consiste en l'application d'une substance chimique sur une peau rasée de lapin pendant 4h et de regarder à 24h si des signes cliniques d'irritations sont présents. Cette méthode critiquable autant d'un point de vue éthique que qualitative reste actuellement le teste le plus utilisé. Depuis le début des années 2000 de nouvelles méthodes in vitro se sont développées tel que le model d'épiderme humain recombiné (RHE). Il s agit d'une multicouche de kératinocyte bien différencié obtenu depuis une culture de don d'ovocyte. Cependant cette méthode en plus d'être très couteuse n'obtient au mieux que 76% de résultat similaire comparé au test in vivo humain. Il existe donc la nécessité de développer une nouvelle méthode in vitro qui simulerait encore mieux la réalité anatomique et physiologique retrouvée in vivo. Notre objectif a été de développer cette nouvelle méthode in vitro. Pour cela nous avons travaillé avec de la peau humaine directement prélevée après une abdominoplastie. Celle ci après préparation avec un dermatome, un couteau dont la lame est réglable pour découper l'épaisseur souhaitée de peau, est montée dans un système de diffusion cellulaire. La couche cornée est alors exposée de manière optimale à 1 ml de la substance chimique testée pendant 4h. L'échantillon de peau est alors fixé dans du formaldéhyde pour permettre la préparation de lames standards d'hématoxyline et éosine. L'irritation est alors investiguée selon des critères histopathologiques de spongioses, de nécroses et de vacuolisations cellulaires. Les résultats de ce.tte première batterie de testes sont plus que prometteurs. En effet, comparé au résultat in vivo, nous obtenons 100% de concordance pour les 4 même substances testes irritantes ou non irritantes, ce qui est supérieur au model d épiderme humain recombiné (76%). De plus le coefficient de variation entre les 3 différentes séries est inférieur à 0.1 ce qui montre une bonne reproductibilité dans un même laboratoire. Dans le futur cette méthode va devoir être testée avec un plus grand nombre de substances chimiques et sa reproductibilité évaluée dans différents laboratoires. Mais cette première evaluation, très encourageante, ouvre des pistes précieuses pour l'avenir des tests irritatifs.

Validation of plant dynamic model by online and laboratory measurements – a tool to predict online COD loads out of production of mechanical printing papers

COD discharges out of processes have increased in line with elevating brightness demands for mechanical pulp and papers. The share of lignin-like substances in COD discharges is on average 75%. In this thesis, a plant dynamic model was created and validated as a means to predict COD loading and discharges out of a mill. The assays were carried out in one paper mill integrate producing mechanical printing papers. The objective in the modeling of plant dynamics was to predict day averages of COD load and discharges out of mills. This means that online data, like 1) the level of large storage towers of pulp and white water 2) pulp dosages, 3) production rates and 4) internal white water flows and discharges were used to create transients into the balances of solids and white water, referred to as “plant dynamics”. A conversion coefficient was verified between TOC and COD. The conversion coefficient was used for predicting the flows from TOC to COD to the waste water treatment plant. The COD load was modeled with similar uncertainty as in reference TOC sampling. The water balance of waste water treatment was validated by the reference concentration of COD. The difference of COD predictions against references was within the same deviation of TOC-predictions. The modeled yield losses and retention values of TOC in pulping and bleaching processes and the modeled fixing of colloidal TOC to solids between the pulping plant and the aeration basin in the waste water treatment plant were similar to references presented in literature. The valid water balances of the waste water treatment plant and the reduction model of lignin-like substances produced a valid prediction of COD discharges out of the mill. A 30% increase in the release of lignin-like substances in the form of production problems was observed in pulping and bleaching processes. The same increase was observed in COD discharges out of waste water treatment. In the prediction of annual COD discharge, it was noticed that the reduction of lignin has a wide deviation from year to year and from one mill to another. This made it difficult to compare the parameters of COD discharges validated in plant dynamic simulation with another mill producing mechanical printing papers. However, a trend of moving from unbleached towards high-brightness TMP in COD discharges was valid.

Special issue on evolving the e-marketing mindset towards the social web model - Editorial Part 1

Con este proyecto editorial nuestro objetivo es promover un campo de investigación clave en la comercialización de hoy, es decir, la evolución de la mentalidad e-marketing hacia el nuevo modelo de web social.

Auger-type granular fertylizer distributor: matemathical model and dynamic simulation

The objective of this study was to model mathematically and to simulate the dynamic behavior of an auger-type fertilizer applicator (AFA) in order to use the variable-rate application (VRA) and reduce the coefficient of variation (CV) of the application, proposing an angular speed controller θ' for the motor drive shaft. The input model was θ' and the response was the fertilizer mass flow, due to the construction, density of fertilizer, fill factor and the end position of the auger. The model was used to simulate a control system in open loop, with an electric drive for AFA using an armature voltage (V A) controller. By introducing a sinusoidal excitation signal in V A with amplitude and delay phase optimized and varying θ' during an operation cycle, it is obtained a reduction of 29.8% in the CV (constant V A) to 11.4%. The development of the mathematical model was a first step towards the introduction of electric drive systems and closed loop control for the implementation of AFA with low CV in VRA.

Effective diffusivity and convective mass transfer coefficient during the drying of bananas

In this article, a methodology is used for the simultaneous determination of the effective diffusivity and the convective mass transfer coefficient in porous solids, which can be considered as an infinite cylinder during drying. Two models are used for optimization and drying simulation: model 1 (constant volume and diffusivity, with equilibrium boundary condition), and model 2 (constant volume and diffusivity with convective boundary condition). Optimization algorithms based on the inverse method were coupled to the analytical solutions, and these solutions can be adjusted to experimental data of the drying kinetics. An application of optimization methodology was made to describe the drying kinetics of whole bananas, using experimental data available in the literature. The statistical indicators enable to affirm that the solution of diffusion equation with convective boundary condition generates results superior than those with the equilibrium boundary condition.

Hydrological simulation using SWAT model in headwater basin in Southeast Brazil

Hydrological models are important tools that have been used in water resource planning and management. Thus, the aim of this work was to calibrate and validate in a daily time scale, the SWAT model (Soil and Water Assessment Tool) to the watershed of the Galo creek , located in Espírito Santo State. To conduct the study we used georeferenced maps of relief, soil type and use, in addition to historical daily time series of basin climate and flow. In modeling were used time series corresponding to the periods Jan 1, 1995 to Dec 31, 2000 and Jan 1, 2001 to Dec 20, 2003 for calibration and validation, respectively. Model performance evaluation was done using the Nash-Sutcliffe coefficient (E NS) and the percentage of bias (P BIAS). SWAT evaluation was also done in the simulation of the following hydrological variables: maximum and minimum annual daily flowsand minimum reference flows, Q90 and Q95, based on mean absolute error. E NS and P BIAS were, respectively, 0.65 and 7.2% and 0.70 and 14.1%, for calibration and validation, indicating a satisfactory performance for the model. SWAT adequately simulated minimum annual daily flow and the reference flows, Q90 and Q95; it was not suitable in the simulation of maximum annual daily flows.

Bartlett-Lewis of rectangular pulse modified model: estimate of parameters for simulation of precipitation in sub-hourly duration

The Bartlett-Lewis Rectangular Pulse Modified (BLPRM) model simulates the precipitous slide in the hourly and sub-hourly and has six parameters for each of the twelve months of the year. This study aimed to evaluate the behavior of precipitation series in the duration of 15 min, obtained by simulation using the model BLPRM in situations: (a) where the parameters are estimated from a combination of statistics, creating five different sets; (b) suitability of the model to generate rain. To adjust the parameters were used rain gauge records of Pelotas/RS/Brazil, which statistics were estimated - mean, variance, covariance, autocorrelation coefficient of lag 1, the proportion of dry days in the period considered. The results showed that the parameters related to the time of onset of precipitation (λ) and intensities (μx) were the most stable and the most unstable were ν parameter, related to rain duration. The BLPRM model adequately represented the mean, variance, and proportion of the dry period of the series of precipitation lasting 15 min and, the time dependence of the heights of rain, represented autocorrelation coefficient of the first retardation was statistically less simulated series suitability for the duration of 15 min.

Turbulent shallow-water model for orographic subgrid-scale perturbations

A parallel pseudo-spectral method for the simulation in distributed memory computers of the shallow-water equations in primitive form was developed and used on the study of turbulent shallow-waters LES models for orographic subgrid-scale perturbations. The main characteristics of the code are: momentum equations integrated in time using an accurate pseudo-spectral technique; Eulerian treatment of advective terms; and parallelization of the code based on a domain decomposition technique. The parallel pseudo-spectral code is efficient on various architectures. It gives high performance onvector computers and good speedup on distributed memory systems. The code is being used for the study of the interaction mechanisms in shallow-water ows with regular as well as random orography with a prescribed spectrum of elevations. Simulations show the evolution of small scale vortical motions from the interaction of the large scale flow and the small-scale orographic perturbations. These interactions transfer energy from the large-scale motions to the small (usually unresolved) scales. The possibility of including the parametrization of this effects in turbulent LES subgrid-stress models for the shallow-water equations is addressed.

A new model for the population pharmacokinetics of didanosine in healthy subjects

Didanosine (ddI) is a component of highly active antiretroviral therapy drug combinations, used especially in resource-limited settings and in zidovudine-resistant patients. The population pharmacokinetics of ddI was evaluated in 48 healthy volunteers enrolled in two bioequivalence studies. These data, along with a set of co-variates, were the subject of a nonlinear mixed-effect modeling analysis using the NONMEM program. A two-compartment model with first order absorption (ADVAN3 TRANS3) was fitted to the serum ddI concentration data. Final pharmacokinetic parameters, expressed as functions of the co-variates gender and creatinine clearance (CL CR), were: oral clearance (CL = 55.1 + 240 x CL CR + 16.6 L/h for males and CL = 55.1 + 240 x CL CR for females), central volume (V2 = 9.8 L), intercompartmental clearance (Q = 40.9 L/h), peripheral volume (V3 = 62.7 + 22.9 L for males and V3 = 62.7 L for females), absorption rate constant (Ka = 1.51/h), and dissolution time of the tablet (D = 0.43 h). The intraindividual (residual) variability expressed as coefficient of variation was 13.0%, whereas the interindividual variability of CL, Q, V3, Ka, and D was 20.1, 75.8, 20.6, 18.9, and 38.2%, respectively. The relatively high (>30%) interindividual variability for some of these parameters, observed under the controlled experimental settings of bioequivalence trials in healthy volunteers, may result from genetic variability of the processes involved in ddI absorption and disposition.

Sleep deprivation reduces the lymphocyte count in a non-obese mouse model of type 1 diabetes mellitus

The objective of the present study was to determine whether sleep deprivation (SD) would promote changes in lymphocyte numbers in a type 1 diabetes model (non-obese diabetic, NOD, mouse strain) and to determine whether SD would affect female and male NOD compared to Swiss mice. The number of lymphocytes in peripheral blood after 24 and 96 h of SD (by multiple platform method) or equivalent period of time in home-cage controls was examined prior to the onset of diabetes. SD for 96 h significantly reduced lymphocytes in male Swiss mice compared to control (8.6 ± 2.1 vs 4.1 ± 0.7 10³/µL; P < 0.02). In male NOD animals, 24- and 96-h SD caused a significant decrease of lymphocytes compared to control (4.4 ± 0.3 vs 1.6 ± 0.5; P < 0.001 and 4.4 ± 0.3 vs 0.9 ± 0.1 10³/µL; P < 0.00001, respectively). Both 24- and 96-h SD induced a reduction in the number of lymphocytes in female Swiss (7.5 ± 0.5 vs 4.5 ± 0.5, 4.4 ± 0.6 10³/µL; P < 0.001, respectively) and NOD mice (4 ± 0.6 vs 1.8 ± 0.2, 1.2 ± 0.4 10³/µL; P < 0.01, respectively) compared to the respective controls. Loss of sleep induced lymphopenia in peripheral blood in both genders and strains used. Since many cases of autoimmunity present reduced numbers of lymphocytes and, in this study, it was more evident in the NOD strain, our results suggest that SD should be considered a risk factor in the onset of autoimmune disorders.

Cloning, sequence analysis, and expression of cDNA coding for the major house dust mite allergen, Der f 1, in Escherichia coli

Our objective was to clone, express and characterize adult Dermatophagoides farinae group 1 (Der f 1) allergens to further produce recombinant allergens for future clinical applications in order to eliminate side reactions from crude extracts of mites. Based on GenBank data, we designed primers and amplified the cDNA fragment coding for Der f 1 by nested-PCR. After purification and recovery, the cDNA fragment was cloned into the pMD19-T vector. The fragment was then sequenced, subcloned into the plasmid pET28a(+), expressed in Escherichia coli BL21 and identified by Western blotting. The cDNA coding for Der f 1 was cloned, sequenced and expressed successfully. Sequence analysis showed the presence of an open reading frame containing 966 bp that encodes a protein of 321 amino acids. Interestingly, homology analysis showed that the Der p 1 shared more than 87% identity in amino acid sequence with Eur m 1 but only 80% with Der f 1. Furthermore, phylogenetic analyses suggested that D. pteronyssinus was evolutionarily closer to Euroglyphus maynei than to D. farinae, even though D. pteronyssinus and D. farinae belong to the same Dermatophagoides genus. A total of three cysteine peptidase active sites were found in the predicted amino acid sequence, including 127-138 (QGGCGSCWAFSG), 267-277 (NYHAVNIVGYG) and 284-303 (YWIVRNSWDTTWGDSGYGYF). Moreover, secondary structure analysis revealed that Der f 1 contained an a helix (33.96%), an extended strand (17.13%), a ß turn (5.61%), and a random coil (43.30%). A simple three-dimensional model of this protein was constructed using a Swiss-model server. The cDNA coding for Der f 1 was cloned, sequenced and expressed successfully. Alignment and phylogenetic analysis suggests that D. pteronyssinus is evolutionarily more similar to E. maynei than to D. farinae.