Using molecular dynamics for the refinement of atomistic models of GPCRs by homology modeling

Despite GPCRs sharing a common seven helix bundle, analysis of the diverse crystallographic structures available reveal specific features that might be relevant for ligand design. Despite the number of crystallographic structures of GPCRs steadily increasing, there are still challenges that hamper the availability of new structures. In the absence of a crystallographic structure, homology modeling remains one of the important techniques for constructing 3D models of proteins. In the present study we investigated the use of molecular dynamics simulations for the refinement of GPCRs models constructed by homology modeling. Specifically, we investigated the relevance of template selection, ligand inclusion as well as the length of the simulation on the quality of the GPCRs models constructed. For this purpose we chose the crystallographic structure of the rat muscarinic M3 receptor as reference and constructed diverse atomistic models by homology modeling, using different templates. Specifically, templates used in the present work include the human muscarinic M2; the more distant human histamine H1 and the even more distant bovine rhodopsin as shown in the GPCRs phylogenetic tree. We also investigated the use or not of a ligand in the refinement process. Hence, we conducted the refinement process of the M3 model using the M2 muscarinic as template with tiotropium or NMS docked in the orthosteric site and compared with the results obtained with a model refined without any ligand bound.     

Effect of zinc nutrition on salinity-induced oxidative damages in wheat genotypes differing in zinc deficiency tolerance

Zinc deficiency and salinity are well-documented soil problems and often occur simultaneously in cultivated soils. Usually, plants respond to environmental stress factors by activating their antioxidative defense mechanisms. The antioxidative response of wheat genotypes to salinity in relation to Zn nutrition is not well understood. So, we investigated the effect of Zn nutrition on the growth, membrane permeability and sulfhydryl group (–SH groups) content of root cells and antioxidative defense mechanisms of wheat plants exposed to salt stress. In a hydroponic experiment, three bread wheat genotypes (Triticum aestivum L. cvs. Rushan, Kavir, and Cross) with different Zn-deficiency tolerance were exposed to adequate (1 μM Zn) and deficient (no Zn) Zn supply and three salinity levels (0, 60, and 120 mM NaCl). The results obtained showed that adequate Zn nutrition counteracted the detrimental effect of 60 mM NaCl level on the growth of all three wheat genotypes while it had no effect on the root and shoot growth of ‘Rushan’ and ‘Kavir’ at the 120 mM NaCl treatment. At the 0 and 60 mM NaCl treatments, Zn application decreased root membrane permeability while increased –SH group content and root activity of catalase (CAT) and superoxide dismutase (SOD) in ‘Rushan’ and ‘Kavir’. In contrast, Zn had no effect on the root membrane permeability and –SH group content of ‘Rushan’ and ‘Kavir’ exposed to the 120 mM NaCl treatment. At all salinity levels, ‘Cross’ plants supplied with Zn had lower root membrane permeability and higher –SH group content compared to those grown under Zn-deficient conditions. At the 0 and 60 salinity levels, Zn-deficient roots of Kavir and Rushan genotype leaked significantly higher amounts of Fe and K than the Zn-sufficient roots. In contrast, at the 120 mM treatment, Zn application had no effect or slightly increased Fe and K concentration in the root ion leakage of these wheat genotypes. For ‘Cross’, at all salinity levels, Zn-deficient roots leaked significantly higher amounts of Fe and K compared with the Zn-sufficient roots. The differential tolerance to salt stress among wheat genotypes examined in this study was related to their tolerance to Zn-deficiency, –SH group content, and root activity of CAT and SOD. Greater tolerance to salinity of Zn-deficiency tolerant genotype ‘Cross’ is probably associated with its greater antioxidative defense capacity.     

Chitosan prevents oxidative stress-induced amyloid β formation and cytotoxicity in NT2 neurons: involvement of transcription factors Nrf2 and NF-κB

Increased oxidative stress is a widely accepted factor in the development and progression of Alzheimer’s disease. Here, we introduce chitosan, an antioxidant oligosaccharide, as a protective agent against H₂O₂/FeSO₄-induced cell death in the NT2 neural cell line. Chitosan not only protects the neurons against cell death, as measured by MTT and caspase-3 activity, but also decreases amyloid β formation. NT2 neurons can be used to elucidate the relationship between oxidative stress and Aβ formation. We induced Aβ formation through oxidative stress in NT2 neurons and studied the effect of chitosan. We demonstrate that chitosan can be neuroprotective by suppressing Aβ formation. We further show that chitosan exerts its protective effect by up-regulation of HO-1, γ-GCS, Hsp-70, and Nrf2, while it inhibits activation of caspase-3 and NF-κB. Chitosan or chitosan derivatives have potential value as neuroprotective agents, particularly with regard to oxidative stress.     

A Carbon Footprint of Textile Recycling: A Case Study in Sweden

Global population growth and rising living standards are increasing apparel consumption. Consequently, consumption of resources and generation of textile waste are increasing. According to the Swedish Environmental Protection Agency, textile consumption increased by 40% between the years 2000 and 2009 in Sweden. Given that there is currently no textile recycling plant in Sweden, the aim of this article is to explore the potential environmental benefits of various textile recycling techniques and thereby direct textile waste management strategies toward more sustainable options. Three different recycling techniques for a model waste consisting of 50% cotton and 50% polyester were identified and a life cycle assessment (LCA) was made to assess the environmental performance of them. The recycling processes are: material reuse of textile waste of adequate quality; separation of cellulose from polyester using N‐methylmorpholine‐N‐oxide as a solvent; and chemical recycling of polyester. These are compared to incineration, representing conventional textile waste treatment in Sweden. The results show that incineration has the highest global warming potential and primary energy usage. The material reuse process exhibits the best performance of the studied systems, with savings of 8 tonnes of carbon dioxide equivalents (CO₂‐eq) and 164 gigajoules (GJ) of primary energy per tonne of textile waste. Sensitivity analyses showed that results are particularly sensitive to the considered yields of the processes and to the choice of replaced products. An integration of these recycling technologies for optimal usage of their different features for treatment of 1 tonne of textile waste shows that 10 tonnes CO₂‐eq and 169 GJ of primary energy could be saved.     

Effect of salinity and waterlogging on growth,anatomical and antioxidative responses in <Emphasis Type="Italic">Mentha aquatica</Emphasis> L.

Salinity and waterlogging are two stresses which in nature often occur simultaneously. In this work, effects of combined waterlogging and salinity stresses are studied on the anatomical alteration, changes of enzymatic antioxidant system and lipid peroxidation in Mentha aquatica L. plants. Seedlings were cultured in half-strength Hoagland medium 50 days after sowing, and were treated under combination of three waterlogging levels (well drained, moderately drained and waterlogging) and NaCl (0, 50, 100, 150 mM) for 30 days. Moderately drained and waterlogging conditions induced differently aerenchyma formation in roots of M. aquatica salt-treated and untreated plants. Moreover, stele diameter and endodermis layer were also affected by salt stress and waterlogging. Salt stress significantly decreased growth, relative water content (RWC), protein level, catalase (CAT) and polyphenol oxidase (PPO) activities, and increased proline content, MDA content, H₂O₂ level and activities of superoxide dismutase (SOD), peroxidase (POX), and ascorbate peroxidase (APX). Waterlogging in salt-untreated plants increased significantly growth parameters, RWC, protein content, antioxidant enzyme activity, and decreased proline content, H₂O₂ and MDA levels. In salt-treated plant, waterlogging caused strong induction of antioxidant enzymes activities especially at severe stress condition. These results suggest M. aquatica is a waterlogging tolerant plant due to significant increase of antioxidant activity, membrane stability and growth under water stress. High antioxidant capacity under waterlogging can be a protective strategy against oxidative damage, and help to salt stress alleviation.     

Determination of clarithromycin in human serum by high-performance liquid chromatography after pre-column derivatization with 9-fluorenylmethyl chloroformate: application to a bioequivalence study

A sensitive liquid chromatographic method for the analysis of clarithromycin, a macrolide antibiotic, in human serum using pre-column derivatization with 9-fluorenylmethyl chloroformate (FMOC-Cl) is described. The method involved liquid-liquid extraction of the drug and an internal standard (amantadine) followed by pre-column derivatization of the analytes with FMOC-Cl. A mixture of 0.05 M phosphate buffer containing triethylamine (2 mL L(-1); pH 3.8) and methanol (17:83, v/v) was used as mobile phase and chromatographic separation was achieved on a Shimpack CLC-ODS column. The eluate was monitored by a fluorescence detector with respective excitation and emission wavelengths of 265 and 315 nm. The analytical method was linear over the concentration range of 0.025-10 microg mL(-1) of clarithromycin in human serum with a limit of quantification of 0.025 microg mL(-1). The assay is sensitive enough to measure drug levels obtained in human single dose studies. In the present method, sensitivity and run time of analysis have been improved, and successfully applied in a bioequivalence study of three different clarithromycin preparations in 12 healthy volunteers.     

Vitamin C counteracts miR-302/367-induced reprogramming of human breast cancer cells and restores their invasive and proliferative capacity

Epigenetic reprogramming by embryonic stem cell-specific miR-302/367 cluster has shown some tumor suppressive effects in cancer cells of different tissues such as skin, colon, and cervix. Vitamin C has been known as a reprogramming enhancer of human and mouse somatic cells. In this study, first we aimed to investigate whether exogenous induction of miR-302/367 in breast cancer cells shows the same tumor suppressive effects previously observed in other cancer cells lines, and whether vitamin C can enhance reprogramming of breast cancer cells and also improve the tumor suppressive function of miR-302/367 cluster. Overexpression of miR-302/367 cluster in MDA-MB-231 and SK-BR-3 breast cancer cells upregulated expression of miR-302/367 members and also some core pluripotency factors including OCT4A, SOX2 and NANOG, induced mesenchymal to epithelial transition, suppressed invasion, proliferation, and induced apoptosis in the both cell lines. However, treatment of the miR-302/367 transfected cells with vitamin C suppressed the expression of pluripotency factors and augmented the tumorigenicity of the breast cancer cells by restoring their proliferative and invasive capacity and compromising the apoptotic effect of miR-302/367. Supplementing the culture medium with vitamin C downregulated expression of TET1 gene which seems to be the reason behind the negative impact of vitamin C on the reprogramming efficiency of miR-302/367 cluster and its anti-tumor effects. Therefore application of vitamin C may not always serve as a reprogramming enhancer depending on its switching function on TET1. This phenomenon should be carefully considered when considering a reprogramming strategy for tumor suppression.     

Molecular investigation of association between common IL-6 polymorphism with cytomegalovirus (CMV) infection and recurrent miscarriage in Iranian women

Molecular Biology Reports - Recurrent pregnancy loss (RPL) is described as two or more spontaneous abortions. To date, scientists in various fields of knowledge, such as genetics, endocrinology,...     

The Role of 5-HT1A Receptors and Neuronal Nitric Oxide Synthase in a Seizur Induced Kindling Model in Rats

Neurochemical Research - Dentate gyrus (DG) has a high density of 5-HT1A receptors. It has neural nitric oxide synthase (nNOS), which is involved in neural excitability. The purpose of this study...     

Co-Flocculation of Yeast Species,a New Mechanism to Govern Population Dynamics in Microbial Ecosystems

Flocculation has primarily been studied as an important technological property of Saccharomyces cerevisiae yeast strains in fermentation processes such as brewing and winemaking. These studies have led to the identification of a group of closely related genes, referred to as the FLO gene family, which controls the flocculation phenotype. All naturally occurring S. cerevisiae strains assessed thus far possess at least four independent copies of structurally similar FLO genes, namely FLO1, FLO5, FLO9 and FLO10. The genes appear to differ primarily by the degree of flocculation induced by their expression. However, the reason for the existence of a large family of very similar genes, all involved in the same phenotype, has remained unclear. In natural ecosystems, and in wine production, S. cerevisiae growth together and competes with a large number of other Saccharomyces and many more non-Saccharomyces yeast species. Our data show that many strains of such wine-related non-Saccharomyces species, some of which have recently attracted significant biotechnological interest as they contribute positively to fermentation and wine character, were able to flocculate efficiently. The data also show that both flocculent and non-flocculent S. cerevisiae strains formed mixed species flocs (a process hereafter referred to as co-flocculation) with some of these non-Saccharomyces yeasts. This ability of yeast strains to impact flocculation behaviour of other species in mixed inocula has not been described previously. Further investigation into the genetic regulation of co-flocculation revealed that different FLO genes impact differently on such adhesion phenotypes, favouring adhesion with some species while excluding other species from such mixed flocs. The data therefore strongly suggest that FLO genes govern the selective association of S. cerevisiae with specific species of non-Saccharomyces yeasts, and may therefore be drivers of ecosystem organisational patterns. Our data provide, for the first time, insights into the role of the FLO gene family beyond intraspecies cellular association, and suggest a wider evolutionary role for the FLO genes. Such a role would explain the evolutionary persistence of a large multigene family of genes with apparently similar function.