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.
Ninety five samples were collected from different samples (urine, ear and wounds swaps), from hospitals in Baghdad city. The results of cultural, microscopic , biochemical tests indicated that in urine samples E.coli have high occurrence frequency 19 (47.5%) followed by Proteus mirabilis 18(45%) and Klebsiella species 1 (2.5%), while in wounds samples each of Pseudomonas spp. and Proteus mirabilis 10 (25%) , then followed by E.coli with 5 (12.5%) and Klebsiella species 3 (7.5%). Ear swaps samples revealed that Pseudomonas aeruginosa 7 (46%) was the major bacterium followed by Proteus mirabilis 4(26.6).Sensitivity test against eleven antimicrobial agents was done for all of the Proteus mirabilis isolates (32 isolates). The results display
... Show MoreThe current study included the separation of three alkaloid compounds from Anastatica Hierochuntica and studied the effect of the these compounds on cancerous cells , specifically liver cancer it was found that compound number one is the most influential or inhibiting at 50 percent followed by compound number three when using concentration of 400 μg/mL.
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Mushrooms have bioactive compounds that have antimicrobial, anti-cancer and antioxidant activities among other medicinal benefits. In the present study, we examined the anti-cell proliferation activities of mushrooms from eight genera obtained from the wild in Nigeria. Saccharomyces cerevisiae was used as a model organism to screen mushroom extracts for anti-cell proliferation activity. Polyphenols, high molecular weight polysaccharides and low molecular weight compounds from aqueous extracts were obtained from the test mushrooms using methanol and water respectively. The extracts were screened in vitro at different concentrations of extracts with the CyQuant cell proliferation assay. The high molecular
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This study was conducted to prepare protein concentrates from AL-Zahdidate’s pits by using alkaline methods where the chemical composition of the pits were (7.30, 1.04, 5.80, 8.68 and 77.19) % for each of the moisture, ash, protein, fat and carbohydrates respectively and the chemical composition of the concentrate protein was (6.62, 4.10, 26.70, 0.93, and 58.65) % respectively. The content of protein concentrate from the metallic elements (144.07, 25.11, 15.02, 0.49, 0.59, 0.27, 0.22 and 234.6) mg/ 100 g each of potassium, magnesium, calcium, iron, manganese, copper, zinc and phosphorus respectively. The results of SDS-PAGE showed five bands with weights molecular ranged between 11000-70000 Dalton. Give the biscuit which contain protei
... Show MoreOrganofluorines, as a pollutant, belongs to a group of substances which are very difficult to neutralize. They are part of many products of everyday use and for this reason they pollute the environment in large quantities. Perfluorinated carboxylic acids are entered into the list of the “Stockholm Convention on Persistent Organic Pollutants” in order to minimize the load on the environment by significantly reducing their use, up to their complete rejection. The DD4 strain was isolated from the soil by the enrichment method and identified using 16S rRNA method as Pseudomonas plecoglossicida. It is able to metabolize perfluorooctanoic acid (PFOA) as the only carbon source in Raymond nutrient medium with a concentration of 1000
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Non-thermal (low-temperature) plasma may act as an alternative approach to control superficial wound and skin infections when the effectiveness of chemical agents is weak due to natural pathogen or biofilm resistance. In this paper an atmospheric pressure plasma needle jet device which generates a cold plasma jet is used to measure the effectiveness of plasma treatment against different pathogenic bacteria and to test the individual susceptibility of pathogenic bacteria to non-thermal argon plasma. It is found that, Gram-negative bacteria were more susceptible to plasma treatment than Gram-positive bacteria. For the Gram-negative bacteria Pseudomonas aeruginosa, there were no survivors among the initial 1x108C.F.U (Co
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