The aim of study To purify GPCR from a local strain of S. cerevisiae using Ion exchange and gel filtration chromatography techniques , by packing materials for columns which will be chosen of low cost comparing to the already used in published researches, which depend on the costly affinity chromatography and other expensive methods of purification. Local strain of S. cerevisiae chosen for extraction and purification of G-protein coupled receptor (GPCR) .The strains were obtained from biology department in Al- Mosul University, Iraq. The isolated colony was activated on Yeast Extract Pepton Dextrose Broth (YEPDB) and incubated at 30 C˚ for 24 h .Loop fully of the yeast culture was transferred to (10ml) of yeast extract peptone glucose agar (YEPGA) slant , then incubated at 30C˚for 24h , after that it was stored at 4C˚ ,the yeast cultures were reactivated and persevered after each two weeks period. S.cerevisiae was identified by morphological, microscopic characterization and biochemical test . The GPCR that extract from whole cell of S.cerevisiae was purified by ion exchange chromatography using DEAE-Sepharose ,the bound proteins (negatively charged) were then eluted using gradient concentration of NaCl ranged between( 0.1 -0.5M). Gel filtration chromatography using Sepharose 6B was applied as a second step of purification. The optical density for each fraction was measured at 280 nm by UV-VS spectrophotometer then the GPCR concentration was determined by using ELISA Kit . The fractions which gave the highest absorbance and concentration of GPCR were collected .The molecular weight of GPCR was determined by gel filtration chromatography using blue dextrin solution. Standard curve was plotted between log of molecular weight for standard protein and the ratio of Ve/Vo of GPCR . The purity of the GPCR that extracted and purified from whole cell of S, cerevisiae were carried out by using SDS-PAGE electrophoresis . In ion exchange chromatography the fraction were collected with 5 ml tube at a flow rate 0.5 ml/ min and eluted with gradient (0.1-0.5M) of sodium chloride solution. Two proteins peaks appeared after eluted by the gradient concentration of sodium chloride, while no protein peaks appeared in the washing fractions. The GPCR concentration was measured in the fractions of these two protein peaks, data indicated that GPCR located in the first protein peak (eluted at 0.1M of NaCl) at fraction numbers between 3 and 9, the maximum concentration of GPCR was 9.281 with specific activity 71.58(ng/mg)protein , 3.125 purification folds and72.9(%) yield while the second peaks (eluted at 0.4 M of NaCl) don't give any concentration for GPCR, thus its neglected. Gel filtration chromatography was used as second step of purification which applied by using sepharose 6B. Results show single active protein peaks appeared that identical with the peak of GPCR at fractions numbers(29-35). The maximum concentration of GPCR was 9.082 (ng/ml)was observed in these fractions. The specific activity for these fractions was 151.37 (ng/mg) protein with 6.608 purification folds and 39.64 (%) yield. The present study a chive a relatively high purification of GPCR from whole cell of a local strain S. cerevisiae with fold purification 6.608 and a yield of 39.64 % and molecular weight about~33KD.
Background: Acne vulgaris is one of the top three most commonly encountered dermatological problems worldwide in both primary and secondary care. Human keratinocytes express functional TLR2 heterodimers. An increased expression of TLR2 was detected in the epidermis of inflammatory acne lesions, as observed in normal skin; the expression level increased with the degree of differentiation of the keratinocytes. TLR2 expression is upregulated in inflammatory acne lesions and induced by C. acnes. The current study conducted to assess the oral isotretinion treatment effect on the acne vulgaris patients by evaluated the Toll Like Receptor 2 as a major immune system marker in Acne vulgaris immune re
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Silicon nanowire arrays (SiNWs) are created utilizing the metal-assisted chemical etching method with an Ag metal as a catalyst and different etching time of 15, 30, and 60 minutes using n-Si (100). Physical properties such as structural, surface morphology, and optical properties of the prepared SiNWs are studied. The diameter of prepared SiNWs ranged from 20 to 280 nm, and the reflectance in the visible part of the wavelength spectrum was less than 1% for all prepared samples. The obtained energy gap of prepared SiNWs was around 2 eV, which is higher than the energy gap of bulk silicon. X-ray diffraction (XRD) has diffraction peaks at 68.70o for all prepared samples. The heterojunction solar cell was fabricated based on the
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Zinc-air fuel cells (ZAFCs) are a promising energy source that could compete with lithium-ion batteries and perhaps proton-exchange membrane fuel cells (PEMFCs) for next-generation electrified transportation and energy storage applications. In the present work, a flow-type ZAFC with mechanical rechargeable was adopted, combined with an auxiliary cell (electrolyzer) for zinc renewal and electrolyte recharge to the main cell. In this work a practical study was performed to calculate the cell capacity (Ah), as well as study the electrolysis cell efficiency by current efficiency, and study the effective parameters that have an influence on cell performance such as space velocity and current density. The best parameters were selected to
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AgInSe2 (AIS) thin films solar cell involving of n-type AgInSe2 and Si of p-type substrate by using thermal evaporation method. The influence of annealing of the preparation AgInSe2 were considered to find the best properties of solar device. Thin film AIS have been deposited under the vacuum of 1.5*10-6 Torr with (400) nm thickness at R.T and annealing temperatures (473,573) K. Polycrystalline tetragonal structure for AIS thin films from XRD and increasing of surface roughness from AFM, energy gap values decreasing with increasing annealing temperatures, all films were negative type, I-V characteristics show increasing of efficiency with increasing of annealing temperatures.
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The aquatic crude extract of Silybum marianum dry grains prepared by melting them in distil water by the method of soak and shake. The effect of Silybum marianum crude extract studied in vitro on three tumor cell line the Hep-2, AMN-3 and RD for 24, 48 and 72 hours of exposure, and one cell line of normal cells REF for 72 hr exposure. The results showed that the prescence of toxic effect of the aquatic crude extract on the cell lines of Hep-2, AMN-3 and RD at 10 and 100 µg/ ml upto the higher concentrations when they exposed to the extract for 48 hr. as compared with the control treatment, and when the exposure period increased to 72 hr. the toxic effect started at low concentrations (5 and 10 µg/ ml) as compared with the control g
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Objective: Per-implantitis is one of the implant treatment complications. Dentists have failed to restore damaged periodontium by using conventional therapies. Tissue engineering (stem cells, scaffold and growth factors) aims to reconstruct natural tissues. The paper aimed to isolate both periodontal ligament stem cells (PDLSCs) and bone marrow mesenchymal stem cells (BMMSCs) and use them in a co-culture method to create three-layered cell sheets for reconstructing natural periodontal ligament (PDL) tissue. Materials and methods: BMMSCs were isolated from rabbit tibia and femur, and PDLSC culture was established from the lower right incisor. The cells were co-cultured to induce BMMSC differentiation into PDL cells. Cell morphology, stem cel
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The effect of using grinded rocks of (quartzite and porcelanite) as powder of (10 and 20) % replacement by weight of cement for self-compacting concrete slabs was investigated in this study. Five slabs with 15 concrete cubes were tested experimentally at 28 days to study the compressive strength, ultimate load, ultimate deflection, ductility, crack load and steel strain. The test results show that, the compressive strength improvement when replacement of local rock powder reached to (7.3, 4.22) % for (10 and 20) % quartzite powder and (11.3, 16.1) % for (10 and 20) % porcelanite powder, respectively compared to the reference specimen. The ultimate load percentage increase for slabs with (10 and 20) % rep
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The focus of this work is on systematically understanding the effects of packing density of the sand grains on both the internal and bulk mechanical properties for strip footing interacting with granular soil. The studies are based on particle image velocimetry (PIV) method, coupled with a high resolution imaging camera. This provides valuable new insights on the evolution of slip planes at grain-scale under different fractions of the ultimate load. Furthermore, the PIV based results are compared with finite element method simulations in which the experimentally characterised parameters and constitutive behaviour are fed as an input, and a good level of agreements are obtained. The reported results would serve to the practicing engineers, r
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