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.
Abstract:
Background: Retinol binding protein 4 (RBP4), an adipokine that participate in a lipid metabolism or insulin resistance through a complex regulatory network. Recently, RBP4 was reported to be associated with many cardiovascular diseases (CVDs) risk factors in patients of type 2 diabetes mellitus (T2DM). This study aims to study the correlation of serum RBP4 with some markers of glycemic control, dyslipidemia, hypertension and obesity in T2DM Iraqi patients.
Subjects and Methods: one hundred fifty participants were enrolled in this coss-sectional study, 120 of participants were T2DM patients and 30 were apparently healthy individuals to serve as control gro
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The current work was designed to investigate serum angiopoietin like protein-8 and hyaluronic acid among Iraqi hemodialysis patients with and without type 2 diabetes mellitus, and to find relationship between them, as well as if these patients are at risk of kidney fibrosis. Subjects & Methods: in this study, serum samples were obtained from (60) Iraqis patients with end stage renal diseases (ESRD)on hemodialysis (HD) (30 patients with T2DM (G2) and 30 patients withoutT2DM (G3)) in addition to (30) healthy individuals as a control group (G1), their ages ranged from (35-65) years. The patients attended the Al-Yarmouk Teaching Hospital, Baghdad. Results: the results in this study showed a highly a significant elevation inserum angiopoietin li
... Show MoreBackground: Ulcerative colitis disease is a chronic inflammatory condition that affects the gastrointestinal tract. In regulation of this inflammatory process, Interleukin-6, C-reactive proteins and albumin have a major role. Overproduction of IL-6 by immunocompetent cells contributes to activate the liver to produce CRP, transudation of plasma albumin and development of the inflammatory condition. Elevated levels of IL-6 in saliva could be expected, because the saliva-producing cells are part of the digestive system. The purpose of this study was to assess salivary IL-6, CRP and albumin in ulcerative colitis patients in relation to oral findings. Materials and methods: Forty eight saliva specimens collected from three groups of subjects (s
... Show MoreBackground: Rheumatoid arthritis is a chronic destructive inflammatory disease associated with destruction of joint connective tissues and bones, affecting 0.5%–1% of the population worldwide reporting higher prevalence of periodontitis among rheumatoid arthritis patients. The purpose of this study is to estimate level of salivary C-reactive protein in relation to the occurrence and severity of the periodontal disease and other oral parameters among group of patients with rheumatoid arthritis Material and methods: Fifty women patients with rheumatoid arthritis; twenty five on Methotrexate treatment and twenty five on combination treatment of Methotrexate and Etanercept selected as study groups with an age range (30-40) years old and
... Show MoreThis study evaluated yellow mealworm (Tenebrio molitor) larvae meal as an alternative animal-protein ingredient in diets for common carp (Cyprinus carpio) fingerlings. Two diets were compared for 60 days: a conventional fish-meal diet (T1; 32.09% crude protein) and a diet in which the animal-protein component was replaced weight-for-weight with T. molitor meal (T2; 23.88% crude protein). Each treatment was tested in duplicate tanks containing 28 fingerlings per tank, and growth was assessed over three consecutive 20-day phases. The initial mean body weight was 3.21 g. Weight gain differed significantly between treatments (p < 0.05), with the highest total value recorded in T2 (4.87 ± 0.47 g). The T2 diet produced a significantly higher fee
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The purpose of this study was to measure serum levels of insulin-like growth factor-binding protein (IGFBP7), Insulin-like Growth Factor 1 (IGF-1), Growth Hormone (GH), Interleukin 6 (IL-6) and insulin in acromegaly patients and healthy controls. The acromegaly group had 60 patients, while the population group had 30 people who had never had acromegaly before. The concentration of IGFBP7, IGF-1, GH, IL-6, and insulin were determined. The results of the present study indicate that IGFBP7 level in the acromegaly group was significantly lower (1.690.07 ng/mL vs. 2.740.12 ng/mL, respectively, p = 0.001). IGF-1, GH, IL-6, and insulin concentrations were also significantly higher in acromegaly patients. The diagnostic accuracy (2.194) was exce
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The purpose of the current investigation was to examine the relationship between inflammation marker C-reactive protein (CRP), an oxidative stress marker (Malondialdehyde; MDA), and DNA damage assessed by comet assay (single-cell gel electrophoresis, SCG) in smokers and non-smokers. The comet assay score was analyzed by image software using three parameters for measuring DNA damage which were tail length, tail DNA %, and tail moment. SCG is commonly used as an in vitro and in vivo genotoxicity test. Because of its ability to detect different forms of DNA damage and its simplicity of application, the technique is being increasingly used in human biomonitoring. The test was performed on blood samples from smokers and non-smokers to bett
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There are several oil reservoirs that had severe from a sudden or gradual decline in their production due to asphaltene precipitation inside these reservoirs. Asphaltene deposition inside oil reservoirs causes damage for permeability and skin factor, wettability alteration of a reservoir, greater drawdown pressure. These adverse changing lead to flow rate reduction, so the economic profit will drop. The aim of this study is using local solvents: reformate, heavy-naphtha and binary of them for dissolving precipitated asphaltene inside the oil reservoir. Three samples of the sand pack had been prepared and mixed with a certain amount of asphaltene. Permeability of these samples calculated before and after mixed with asphaltenes. Then, the
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In this research, carbon nanotubes (CNTs) is prepared through the Hummers method with a slight change in some of the work steps, thus, a new method has been created for preparing carbon nanotubes which is similar to the original Hummers method that is used to prepare graphene oxide. Then, the suspension carbon nanotubes is transferred to a simple electrode position platform consisting of two electrodes and the cell body for the coating and reduction of the carbon nanotubes on ITO glass which represents the cathode electrode while platinum represents the anode electrode. The deposited layer of carbon nanotubes is examined through the scanning electron microscope technique (SEM), and the images throughout the research show the
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