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Research in Pharmaceutical Sciences، جلد ۱۳، شماره ۵، صفحات ۴۱۳-۴۲۱
عنوان فارسی
چکیده فارسی مقاله
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عنوان انگلیسی Effect of buffer additives on solubilization and refolding of reteplase inclusion bodies
چکیده انگلیسی مقاله Reteplase is a non-glycosylated and recombinant form of tissue type plasminogen activator, which is produced in Escherichia coli. However, its overexpression usually leads to formation of inactive aggregates or inclusion bodies. In the present study, we report on the development of optimized processes for isolation, solubilization, and refolding of reteplase inclusion bodies to recover active protein. After protein overexpression in E. coli BL21 (DE3) inclusion bodies were isolated by cell disruption and repeated wash of pellet with buffer containing Triton X-100. To solubilize the inclusion bodies, different types, concentrations, pHs, and additives of denaturing agents were used. Rapid micro dilution method was applied for refolding of solubilized reteplase. Different chemical additives including sugars, alcohols, polymers, detergents, amino acids, kosmotropic, and chaotropic salts, reducing agents, and buffering agents were used in the refolding buffer. To evaluate the biological activity of refolded reteplase, an indirect chromogenic assay was performed. The best solubilizing agent for dissolving reteplase inclusion bodies was 6 M urea at pH 12. The optimized buffer for refolding of solubilized reteplase was found to be 1.15 M glucose, 9.16 mM imidazole, and 0.16 M sorbitol which resulted in high yield of biologically active protein. Our results indicate type, concentration, and pH of solvent and type, concentration, and combination of chemical additives can significantly influence the yield of inclusion bodies solubilization and refolding.
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نویسندگان مقاله | Iman Esmaili
Department of Pharmaceutical Biotechnology and Isfahan Pharmaceutical Research Center, School of Pharmacy and Pharmaceutical Sciences, Isfahan University of Medical Sciences, Isfahan, I.R. Iran.


| Hamid Mir Mohammad Sadeghi
Department of Pharmaceutical Biotechnology and Isfahan Pharmaceutical Research Center, School of Pharmacy and Pharmaceutical Sciences, Isfahan University of Medical Sciences, Isfahan, I.R. Iran.


| Vajihe Akbari


نشانی اینترنتی http://rps.mui.ac.ir/index.php/jrps/article/view/1851
فایل مقاله اشکال در دسترسی به فایل - ./files/site1/rds_journals/115/article-115-756703.pdf
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زبان مقاله منتشر شده en
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نوع مقاله منتشر شده Original Article
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