docهمسانه سازی جایگاه کاتالیتیک اگزوتوکسین A سودوموناس آیروژینوزا
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همسانه سازی جایگاه کاتالیتیک اگزوتوکسین A سودوموناس آیروژینوزا
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Pro Expr and Puri. 2005; 44: 8- Wedekind1 JE, Trame1 CB, Dorywalska M, et 52-7. al. 14- Tramper-Stranders GA, Wolfs TFW, van der Refined crystallographic structure of exotoxin A (PE38KDEL) in íîƱŚŤƀŝŚţìæƽƵŹŚưƃ æíƽƵŹƹŵƱŚŬƳŻƾƳŚƯŹŵƾŤƃřŶƸŝšŚƯŶųƹƾƨƃżěƭƺƬƗƵŚĮƄƳřŵƾƄƷƹĦěƾưƬƗƽ ƶƬŬƯ Downloaded from zums.ac.ir at 17:06 IRST on Tuesday March 7th 2017 èç ƱřŹŚƨưƷƹƾƴǀƯřƭřźƸŝ Ent CK. Detection of Pseudomonas aeruginosa in aeruginosa exotoxin A. J biolo Biochem. 2002; patients with cystic fibrosis. J Cystic Fibrosis. 277: 46669-75. 2005; 4: 37-43. 18- Bjon MJ, Iglewskl BH, Ives SK, Sadoff JC, 15- Xiao X, Zhang J, Gong J, et al. Rapid Vasll ML. Effect of iron on yields of exotoxin A detection of Pseudomonas aeruginosa by the in cultures of Pseudomonas aeruginosa PA-103. J fluorescence quantitative TaqMan PCR assay Infec Immun. 1978; 19: 785-91. targeting ETA gene. Chin J Biotech. 2008, 24: 19- Keyvani Amineh H, Ghasemian Safaei H. 581-5. Purification of exotoxin A from Pseudomonas 16- Yates SP, Taylor PL, Jorgensen R, et al. aeruginosa J Tehran Vet Med. 2000; 56:33-6. Structure–function water-soluble 20- Joshi BH, Puri RK. Optimization of inhibitors of the catalytic domain of exotoxin A expression and purification of two biologically from Pseudomonas aeruginosa. J Biochem.2005; active chimeric fusion proteins that consist of 385: 667-75. human interleukin-13 and Pseudomonas exotoxin 17- Armstrong S, Yates SP, Merrill A. Insight into in Escherichia coli. Pro Expr and Puri. 2005; 39: the 189-98. catalytic analysis mechanism of of Pseudomonas íîƱŚŤƀŝŚţìæƽƵŹŚưƃ æíƽƵŹƹŵƱŚŬƳŻƾƳŚ ƯŹŵƾŤƃřŶƸŝšŚƯŶųƹƾƨƃżěƭƺƬƗƵŚĮƄƳřŵƾƄƷƹĦěƾưƬƗƽƶƬŬƯ ƦǀŤǀƫŚţŚƧƵŚĮƿŚūƽŻŚſƶƳŚƀưƷ èè Cloning of Catalytic Domain of Exotoxin A from Pseudomonas Aeruginosa Amini B1, Kamali M2, Zarei Mahmod abadi A3, Mortazavi Y4, Ebrahim habibi A5, Bayat E1, Farhadi N2, Javadi HR2, kyhan AH2 1 Downloaded from zums.ac.ir at 17:06 IRST on Tuesday March 7th 2017 2 Science and Research Branch, Islamic Azad University, Tehran, Iran Nanobiotechnology Research Center, Baqyiatallah University of Medical Sciences, Tehran, Iran 3 Dept. of Biochemistry, Baqyiatallah University of Medical Sciences, Tehran, Iran 4 Dept. of Molecular Medicine and Genetics, Zanjan University of Medical Sciences, Zanjan, Iran 5 Dept. of Endocrinology and Metabolism Research Center, Tehran University of Medical Sciences, Tehran, Iran Corresponding Author: Kamali M, Nanobiotechnology Research Center, Baqyiatallah University of Medical Sciences, Tehran, Iran E-mail: [email protected] Received: 17 Jan 2010 Accepted: 8 Mar 2010 Background and Objective: Antibody against Pseudomonas aeruginosa exotoxin A can be used in immunotherapy together with antibiotics to treat acute burn patients. Exotoxin A is one of the virulence factors in Pseudomonas aeruginosa that comprises of three domains, binding domain, translocation and catalytic domain. The purpose of this study was to construct the recombinant domain of the catalytic part of this microorganism in order to produce antibody against it. Methods and Materials: Pseudomonas aeruginosa samples were isolated from burn patients hospitalized in Mousavi Hospital, Zanjan, Iran and its species was identified by Biochemical tests. Bacteria genomic DNA and also the catalytic domain of exotoxin A was amplified by PCR. PCR products and plasmid extracts was digested by restriction enzymes. Subsequently PCR products and plasmids transformed into E. coli BL21 (DE3). Clones containing gene of interest was determined by restriction enzyme digestion and sequencing. Results: The sequence homology of the catalytic domain of exotoxin A was compared with that of the published gene data banks. The results showed a complete homology between our gene species and the published genome in data banks. Conclusion: The results of this study showed that about 90% of the isolated bacteria contained exotoxin A and there was a sequence homology between our species and published gene data banks. Key word: Pseudomonas aeruginosa, Cloning, Exotoxin A, Catalytic domain íîƱŚŤƀŝŚţìæƽƵŹŚưƃ æíƽƵŹƹŵƱŚŬƳŻƾƳŚƯŹŵƾŤƃřŶƸŝšŚƯŶųƹƾƨƃżěƭƺƬƗƵŚĮƄƳřŵƾƄƷƹĦěƾưƬƗƽ ƶƬŬƯ
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