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: Melanin is a high-molecular weight pigment produced through the oxidative polymerization of phenolic or indolic compounds and plays a perfect role in UV-light shielding, as well as in photoprotection. Among biopolymers, melanin is unique in many aspects. This study is designed to screen Production, extraction and characterizes of an extracellular melanin pigment from clinically isolated P. aeruginosa. Objective: The aim of the current study is isolation and diagnosis of P.aeruginosa using vitek-2 compact system and screening the ability to produce melanin and characterization of extracted melanin by UV-vis, FTIR, XRD and SEM. Materials and methods: the samples swab inoculated on cetrimide agar as selective media and incubated
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Specific microorganisms can produce bacterial nanocellulose (BNC), with acetic acid bacteria (AAB) being the most active producer. The family Acetobacteraceae includes the obligate aerobic, motile acetic acid bacteria. The BNC has attracted a lot of interest across a wide range of industries, including pharmaceuticals, due to its flexible characteristics, properties, and advantages. The present study was conducted to purify and characterize BNC produced from AAB isolated from apple vinegar. Bacterial nanocellulose was synthesized using a natural date palm liquid medium at pH 6 at 30°C for 8–10 days. The bacterial cellulose produced was then purified using a technique involving 0.1 M sodium hydroxide. To ascertain the surface mor
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Chemiluminescenc (CL), light emitted during chemical reaction, is one of the accurate methods used to detect directly oxygen free radicals. In this study, luminol was used as CL detector, to detect the concentration of free radicals formed in whole blood exposed to high power microwave pulses. The changes in the intensity of CL signal gives a clear relation between the concentration of free radicals formed by radiation in blood and changes in blood properties such as hemolysis of blood cells. This is done by measuring the electrical sytoplsimic electrical properties, the results are substituted in Maxwell-Wagner equation, to obtain electrical conductivity of cytoplasm, which is 18.3 ms/cm, while at suspension med
... Show MoreThis study was conducted to isolate and identify killer yeast Hanseniaspora uvarum from dates vinegar and measurement the ability of this yeast to produce killer toxin. The antimicrobial activity of the concentrated supernatant containing partially purified concentrated killer toxin was also detected against several pathogenic bacteria and yeast species, which includes two types of yeast Rhodotorula mucilaginosa and Candida tropicalis and four human pathogenic bacteria Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeurginosa. In addition, the antagonistic activity of examined yeast have been studied toward four types of fungi, where two are pathogenic
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An increasing trend to use probiotic and study their effects on the pathogens has been conductor where they are defined as live micro-organisms that give a health benefit to the host when ingested in sufficient quantities, including the yeast Saccharomyces boulardii In addition research show that a magnetic field (MF) has a biological effect. This study aims to investigate the effects of magnetic field on the inhibitory action of Saccharomyces boulardii against bacteria isolated from urinary tract infection, Study the sensitivity of bacterial isolates to antibiotics after diagnosis by microscopic, Cultural and biochemical examinations as well as Api20 E examinations were used gram negative bacteria , Most isolates were resistant to an
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Klebsiella pneumoniae are Gram-negative which cause many diseases such as urinary tract infections, respiratory tract infections and septicemia. Inulinase is an enzyme used in food manufacture and pharmaceuticals. Inulinase is used in decreasing lipid ratio and, cholesterol in blood and considered as a prebiotic factor inside intestine. Many microorganisms can produce inulinase, such as yeast, fungi and bacteria; among such bacteria: Bacillus spp., Arthrobacter spp., and Pseudomonas spp. but there are no studies about inulinase production by K. pneumoniae have been reported. So the current study aims at investing the ability of producing and purification inulinase by K. pneumoniae. Method: K. pneumoniae were isolated from many hospitals and
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Peroxidase is a class of oxidation-reduction reaction enzyme that is useful for accelerating many oxidative reactions that protect cells from the harmful effects of free radicals. Peroxidase is found in many common sources like plants, animals and microbes and have extensive uses in numerous industries such as industrial, medical and food processing. In this study, P. aeruginosa was harvested to utilize and study its peroxidases. P. aeruginosa was isolated from a burn patient, and the isolate was verified as P. aeruginosa using staining techniques, biochemical assay, morphological, and a sensitivity test. The gram stain and biochemical test result show rod pink gram-ne
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General Background: Breast cancer is the most prevalent cancer affecting women, with increasing incidence worldwide. Specific Background: Recent research has focused on the role of epigenetic changes in DNA damage, repair mechanisms, and the potential therapeutic effects of probiotics. Probiotics have shown promise in promoting tissue regeneration and DNA repair. Knowledge Gap: However, the precise impact of probiotics on DNA repair in cancer cells, specifically breast cancer cells, remains underexplored. Aims: This study aimed to evaluate the effects of probiotics on DNA damage repair in AMJ13 Iraqi breast cancer cells and assess the cytotoxic effects of probiotics on these cells. Results: Using the comet assay, we found significan
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