Characterization of the bioactive form and molecular determinants of recognition of cyclic enkephalin peptides at the δ-opioid receptor

An extensive and systematic search strategy to determine the conformational profile of 12 cyclic disulfide-bridged opioid peptides with varying affinities at the δ receptor has been carried out to identify the structure that is recognized by the δ receptor for each analogue. The methods and procedures used here for the conformational search have already been validated for [D -Pen2, D -Pen5] enkephalin (DPDPE), one member of this family. Use of these methods led to a low-energy solution conformation of DPDPE in excellent agreement with all the geometric properties deduced from its solution nmr spectra. Each of the analogue was subjected to the same procedure, involving a combination of molecular dynamics simulations at high and low temperature. The study was repeated in two environmental conditions, an apolar environment, simulated by using a distance-dependent dielectric constant, and a polar environment by embedding the peptides in a high constant dielectric ( ε = 80). An automated comparison of the different conformers based on their backbone rms and average distance between the key aromatic moieties was followed by graphic analysis using maximum structural overlap. The cross-comparison of the conformations for each analogue revealed a unique conformer that may be recognized by the δ receptor for each high-affinity analogue that permitted maintaining the critical elements required for recognition in a simple spatial orientation, while maximizing similarity in other regions. © 1993 John Wiley & Sons, Inc.     

A case study of airborne culturable microorganisms in a poultry slaughterhouse in Styria, Austria

According to Council Directive 90/679/EEC on the protection of workers from risks related to exposure to biological agents at work, nature, degree and duration of workers’ exposure to microorganisms must be determined. This directive has already been implemented in waste and wastewater management. The present case study investigates concentration and composition of microorganisms in a poultry slaughterhouse in the State of Styria, Austria. From June to November 2002, measurements were conducted at the sampling sites ‘moving rail’ and ‘gall bladder separation’ using the Andersen six stage viable cascade impactor and the SKC BioSampler. The results of this study were compared with other previous studies which were carried out using the same device (ACFM) and the same measurement methods. At the processing area of the ‘moving rail’, the median concentration of airborne mesophilic bacteria was 1.7×10⁶ CFU/m³ which is 8000 times higher than the background concentration of residential areas (approx. 210 CFU/m³). The airborne microorganisms concentration was 1.7×10⁴ CFU/m³ at composting plants which is 100 times lower than at a workplace of a poultry slaughterhouse. The study shows that poultry slaughterhouse employees are exposed to high concentrations of airborne microorganisms throughout the entire work time without using a respiratory protective device. For the protection of employees against airborne biological agents, relevant measures should be introduced to this field of work.     

Multiple Caretaking of Efe (Pygmy) Infants

Two models of the human infant's caretaking requirements—the continuous care and contact model and the caretaker-child strategy model—are discussed in terms of the caretaking practices observed among the Efe (Pygmies) of northeastern Zaire. The Efe¹ engage in a system of multiple care which begins at birth and continues through at least the first 18 weeks of life. An important aspect of this care includes being suckled by lactating and nonlactating women. The data suggest that the continuous care and contact model is too rigid a formulation of the infant's caretaking environment, and lend support to the strategic model. Efe caretaking practices are discussed in terms of the cultural, ecological, and physiological constraints acting on these people.     

Inhibitory Potential of a Designed Peptide Inhibitor Based on Zymogen Structure of Trypsin from Spodoptera frugiperda: In Silico Insights

Spodoptera frugiperda (J.E. Smith) is an invasive pest in agriculture. It can potentially damage yield resulting severe crop losses and subsequently significant economic damage each year. S. frugiperda is predominantly managed using traditional chemical pesticides. Accordingly, sustainable alternatives such as digestive enzymes inhibitors can be used as an efficient pest management that protects the environment. This contribution aims to examine the pro-region of S. frugiperda trypsin as specific inhibitor of the pest protease enzyme. Structural modeling in conjunction with molecular docking simulations were conducted to design a peptide sequence with the best docking scores and strong binding energy to the target enzyme. The structural models of six pro-peptides were produced based on modification of 7-amino acids of the pro-region of S. frugiperda trypsin. VERIFY_3D, ERRAT, PROCHECK, PROSA and WHAT-IF scores validated the reliability of the predicted model of S. frugiperda trypsin. Molecular docking studies between the six designed inhibitor peptides and the predicted model structure at three different pH conditions were carried out. Data revealed that VPSNPQR at pH 11.0 with the best docking score, the lowest binding energy (ΔG) and dissociation constant (K_d) indicated a potent binding affinity towards S. frugiperda trypsin’s active site. Moreover, the peptide showed a weak potential for interaction with the human trypsin. The results indicated the importance of computational studies in design and selection of inhibitor peptides against target enzymes. Such inhibitors can be used for S. frugiperda control, which can be further applied in other pest management programs.

Graphical Abstract

Docking simulations between the pro-peptide inhibitor and Spodoptera frugiperda midgut trypsin confirmed the capacity of the designed pro-region in inhibiting the insect trypsin.

     

Structure-activity studies of β-carboline analogs

A number of β-carboline analogs have been obtained or synthesized, and their $\text{in vitro}$ receptor affinities and $\text{in vivo}$ antagonist activities determined. The choice of analogs was made in order to explore the importance of the N₉-H, the aromatic nitrogen and the C₃-ester moiety for high-receptor affinity and antagonist activity of this class of benzodiazepine antagonist. Among the analogs investigated, we describe the properties of 3-cyano-β-carboline ($\text{lh}$), the first potent β-carboline antagonist without a carbonyl at the C₃-position.The results obtained indicate: (1) Specific interactions of the C₃-substituent with key cationic receptor sites rather than electron-withdrawing properties are important for high-receptor affinity and antagonist activity. (2) Specific in-plane interactions of the atomatic nitrogen with a cationic receptor site, rather than stacking with neutral aromatic residues of the receptor are also important for high affinity and antagonist activity. (3) While the presence of an N₉H enhances receptor affinity, interaction with an anionic receptor site does not appear essential for antagonist activity.     

Evaluation of corneal structures in myopic eyes more than twenty‐two years after photorefractive keratectomy

The aim of this study is to evaluate corneal epithelial thickness (CET), corneal densitometry (CD) in 84 myopic eyes (57 patients) more than 22 years after photorefractive keratectomy, using anterior segment‐optical coherence tomography (AS‐OCT) and Scheimpflug imaging system. The CET was significantly higher in all operated eyes than in unoperated eyes in central sector. A statistically significant increase in CD in corneal anterior layer of central sector was shown in groups of operated eyes with greater ablation depth respect to unoperated eyes. While there was no significant difference in CD between the operated eyes groups with lower ablation depth and unoperated eyes. A significant trend toward higher values in anterior CD with deeper ablations in central sector was found. These noninvasive imaging techniques allow to better understand the corneal remodeling process after photoablation and to monitor the patients over time.     

MYC Promotes Bone Marrow Stem Cell Dysfunction in Fanconi Anemia

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Quantum chemical studies of methylbenz[a]anthracenes: Metabolism and correlations with carcinogenicity

In the context of the bay region, K-region and radical cation hypotheses for polycyclic aromatic carcinogens, molecular properties were calculated for fourteen methyl derivatives of benz[a]anthracene (BA) related to (1) intrinsic substrate reactivities towards activating and detoxifying metabolism and (2)-the stabilities of the putative carbocation ultimate carcinogens. Allvalence electron methods were used, avoiding the inherent difficulties found in the π-electron methods. The calculated substrate reactivities were found to predict major metabolites sucessfully, supporting the validity of their use in attempted correlations with observed carcinogenic potencies. Positive correlations were found between observed carcinogenic potencies and (1) the reactivities of the parent polycyclic aromatic hydrocarbons (PAH) towards the initial distal bay region epoxidation and (2) the stabilities of the diol epoxide carbocations. This latter correlation holds when both the methyl derivatives of BA and previously studied unsubstituted PAH are considered together, indicating its potential usefulness as a screening parameter for carcinogenic activity.     

Application of the direct beta counter Matrix 96 for cytotoxic assays: Simultaneous processing and reading of 96 wells using a51Cr-retention assay

To assess the cytotoxic activity of immune cells, we have developed a⁵¹Cr-retention assay in which the radioactivity retained by⁵¹Cr-labeled target cells, following coincubation with cytotoxic cells, is monitored using the automated Matrix 96 beta counter. The Matrix 96 is designed for simultaneously counting 96 samples isolated from a 96-well microplate. It uses 96 uniform and independent detectors operating on the principle of avalanche gas ionization in the Geiger-Muller mode. Samples must be dry because the detectors are of the open-window type. Therefore, samples from the 96 wells of the microplate are simultaneously harvested onto a filter using the MicroMate 196, a 96-well cell harvester, dried and quantified in the Matrix 96. Usually the⁵¹Cr isotope is measured by the detection of gamma radiation in gamma counters. The Matrix 96, however, monitors Auger electrons, which are also emitted by⁵¹Cr. We have shown that the retention assay can be used to monitor the cytotoxic activity of activated lymphocytes including lymphokine-activated killer cells and tumor-infiltrating lymphocytes against various tumor cell lines. This assay is most suitable for experiments in which low E/T ratios are sufficient to detect highly cytotoxic cells, such as clone screening in cloning assays or in limiting-dilution analysis assays. These assays involve processing and reading large numbers of microplates. In this case, the retention assay monitored in the Matrix 96 will improve the work flow and decrease the amount of radioactive waste.This work was supported by the American Cancer Society grant IN-162-C