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 Fifty extremely halophilic bacteria were isolated from local high salient soils named Al-Massab Al-Aam in south of iraq and were identified by using numerical taxonomy. Fourty strains were belong to the genus Halobacterium which included Hb. halobium (10%). Hb. salinarium (12.5%), Hb.cutirubrum (17.5%), Hb-saccharovorum (12.5%), Hb. valismortis (10%) and Hb. volcanii (37.5%). Growth curves were determined. Generation time (hr) in complex media and logarithmic phase were measured and found to be 10.37±0.59 for Hb. salinarium. 6.49 ± 0.24 for Hb.cutirubrum. 6.70±0.48 for Hb-valismonis, and 11.24 ± 0.96 for Hb. volcanii
Due to wind wave actions, ships impacts, high-speed vehicles and others resources of loading, structures such as high buildings rise bridge and electric transmission towers undergo significant coupled moment loads. In this study, the effect of increasing the value of coupled moment and increasing the rigidity of raft footing on the horizontal deflection by using 3-D finite element using ABAQUS program. The results showed that the increasing the coupled moment value leads to an increase in lateral deflection and increase in the rotational angle (α◦). The rotational angle increases from (0.014, 0.15 to 0.19) at coupled moment (120 kN.m), (0.29, 0.31 and 0.49) at coupled moment (240 kN.m) and (0.57, 0.63 and 1.03) at cou
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In this work, we studied the effect of power variation on inductively coupled plasma parameters using numerical simulation. Different values were used for input power (750 W-1500 W), gas temperature 300K, gas pressure (0.02torr), 5 tourns of the copper coil and the plasma was produced at radio frequency (RF) 13.56 MHZ on the coil above the quartz chamber. For the previous purpose, a computer simulation in two dimensions axisymmetric, based on finite element method, was implemented for argon plasma. Based on the results we were able to obtain plasma with a higher density, which was represented by obtaining the plasma parameters (electron density, electric potential, total power, number density of argon ions, el
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Separation of Trigonelline, the major alkaloid in fenugreek seeds, is difficult because the extract of these seeds usually contains Trigonelline, choline, mucilage, and steroidal saponins, in addition to some other substances. This study amis to isolate the quaternary ammonium alkaloid (Trigonelline) and choline from fenugreek seeds (Trigonella-foenum graecum L.) which have similar physiochemical properties by modifying of the classical method. Seeds were defatted and then extracted with methanol. The presence of alkaloids was detected by using Mayer's and Dragendorff's reagents. In this work, trigonilline was isolated with traces of choline by subsequent processes of purification using analytical and preparative TLC techniques.
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This study was conducted in the specialized center of endocrinology and diabetic (AlKindy hospital from June 2004 to April 2005). Sera of 80 women (include 40 diagnosed Hyperprolactinemia (HPro) and 40 healthy women as control) were used to estimate some biochemical parameters which include prolactin (PRL),total fucose (TF), total protein (TP) and protein bound fucose (PBF), and protein bound hexose (PBHex), also TP TF , TP PBF and TP PBHex ratios. A significant elevation in TP and PBHex, in sera of HPro patients compared to control was found while PBHex/TP ratio showed a slight non significant increase in sera of patients compared to control. On the other hand a significant decrease in TF, TF/TP and PBF/TP in ser
... Show MoreVitamin D receptor (VDR) is a nuclear transcription factor that controls gene expression. Its impaired expression was found to be related to different diseases. VDR also acts as a regulator of different pathways including differentiation, inflammation, calcium and phosphate absorption, etc. but there is no sufficient knowledge about the regulation of the gene itself. Therefore, a better understanding of the genetic and epigenetic factors regulating the VDR may facilitate the improvement of strategies for the prevention and treatment of diseases associated with dysregulation of VDR. In the present investigation, a set of databases and methods were used to identify putative functional elements in the VDR locus. Histone modifications, CpG I
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Utilization of bacterial activity for decolorization of coloured products is one the most promising industrial strategy, as an eco-sustainable and cost-competitive alter-native to physicochemical methods. Laccase production from Bacillus sp. was stud-ied for its decolorization influences on different dyes (Indian ink, Brilliant green, Bromothymol blue, Crystal violet, Safranin, Bromophenol blue, Methelen blue, Giemsa stain, Nigrosin, Toluidin blue, Neutral red, Phenol red, Hanna, Blood, Ben-gal rose B, Bromkresol green, 4-Bromoaniline, Aniline blue, 2,6-Dichlorophenol in-vophenol, Curcumin, Acridine orange, Indigo carmine, Xylene cynol FF,10G, Aliza-rine yellow GG and Kongorose). After 5 days of incubation of the spore-bound lac-case wit
... Show MoreA new class of generalized open sets in a topological space, called G-open sets, is introduced and studied. This class contains all semi-open, preopen, b-open and semi-preopen sets. It is proved that the topology generated by G-open sets contains the topology generated by preopen,b-open and semi-preopen sets respectively.
Let be a metric space and be a continuous map. The notion of the -average shadowing property ( ASP ) for a continuous map on –space is introduced and the relation between the ASP and average shadowing property(ASP)is investigated. We show that if has ASP, then has ASP for every . We prove that if a map be pseudo-equivariant with dense set of periodic points and has the ASP, then is weakly mixing. We also show that if is a expansive pseudo-equivariant homeomorphism that has the ASP and is topologically mixing, then has a -specification. We obtained that the identity map on has the ASP if and only if th
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