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.
Introduction and Aim: Klebsiella pneumoniae is a Gram-negative bacterium responsible for a wide range of infections, including respiratory tract infections (RTIs). This research was aimed to study the antibacterial and anti-biofilm effect of AgNPs produced by Gram positive and negative bacteria on RTIs associated with K. pneumoniae. Materials and Methods: The biofilm formation of K. pneumoniae was determined by tube method qualitatively from select bacterial species characterized by UV-Visible spectroscopy. The antibacterial susceptibility of the bacteria AgNPs was tested for their antibacterial and antibiofilm activity on a clinical isolate of K. pneumoniae. Results: K. pneumoniae isolated from RTIs were strong biofilm prod
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Introduction and Aim: Klebsiella pneumoniae is a Gram-negative bacterium responsible for a wide range of infections, including respiratory tract infections (RTIs). This research was aimed to study the antibacterial and antibiofilm effect of AgNPs produced by Gram positive and negative bacteria on RTIs associated with K. pneumoniae. Materials and Methods: The biofilm formation of K. pneumoniae was determined by tube method qualitatively from select bacterial species characterized by UV-Visible spectroscopy. The antibacterial susceptibility of the bacteria AgNPs was tested for their antibacterial and antibiofilm activity on a clinical isolate of K. pneumoniae. Results: K. pneumoniae isolated from RTIs were strong biofilm producers. The ant
... Show MorePetroleum is one of the most important substances consumed by man at present times, a major energy source in this century, petroleum oils can cause environmental pollution during various stages of production, transportation, refining and use, petroleum hydrocarbons pollutions ranging from soil, ground water to marine environment, become an inevitable problem in the modern life, current study focused on bioremediation process of hydrocarbons contaminants that remaining in the bottom of gas cylinders and discharged to the soil. Twenty-four bacterial isolates were isolated from contaminated soils all of them gram negative bacteria, bacterial isolates screening to investigate the ability of biodegradation of hydrocarbons, these isolates inocula
... Show MoreThe present study aims to detect CTX-M-type ESBL from Escherichia coli clinical isolates and to analyze their antibotic susceptibility patterns. One hundred of E. coli isolates were collected from different clinical samples from a tertiary hospital. ESBL positivity was determined by the disk diffusion method. PCR used for amplification of CTX-M-type ESBL produced by E. coli. Out of 100 E. coli isolates, twenty-four isolates (24%) were ESBL-producers. E. coli isolated from pus was the most frequent clinical specimen that produced ESBL (41.66%) followed by urine (34.21%), respiratory (22.23%), and blood (19.05%). After PCR amplification of these 24 isolates, 10 (41.66%) isolates were found to possess CTX-M genes. The CTX-M type ESBL
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Samples of the green algae were collected from water of Shatt al-Arab in Garmat Ali in Basra. After purification, the green algae identified on Enteromorpha sp. The samples were dried and milled, then sulfated polysaccharides were extracted with hot water at 90°C precipitated with absolute ethanol, dialysed and lyophilized. The chemical composition was total sugars 56.4%, protein 1.3% and sulfur 19.7%. Antioxidation activity of sulfated polysaccharides was studied by four method and included estimation of ability of scavenging hydroxylated radicals, the results showed an increased in ability with increasing concentrations. Ability of scavenging and was 59.86% at the concentration of 2.5 mg/ ml, but BHT was 81.36%. Ability of scavenging
... Show MoreIn this study, a novel application of lab-scale dual chambered air-cathode microbial fuel cell (MFC) has been developed for simultaneous bio-treatment of real pharmaceutical wastewater and renewable electricity generation. The microbial fuel cell (MFC) was provided with zeolite-packed anodic compartment and a cation exchange membrane (CEM) to separate the anode and cathode. The performance of the proposed MFC was evaluated in terms of COD removal and power generation based on the activity of the bacterial consortium in the biofilm mobilized on zeolite bearer. The MFC was fueled with real pharmaceutical wastewater having an initial COD concentration equal to 800 mg/L and inoculated with anaerobic aged sludge. Results demo
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The aim of this research is to benefit from recycl the aircraft waste oils which is discarded in sewage network, to be used in preparation of greases for industrial purposes and to reduce the environmental pollution. In this research synthetic greases were prepared with special specifications by mixing the waste oils after treating with (silica gel as adsorbent agent, and filtration to precipitate impurities then heated to 110 C? to get rid of water) bentonite produced in Iraq which is available and cheap with existence of high density polyethylene at specific conditions of ( heating and mixing) . The best weight proportion were reached, then paraffin wax and additives were added to improve the properties of grease and give the
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