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MedLine Citation:
PMID:  23768109     Owner:  NLM     Status:  In-Data-Review    
Reteplase is the recombinant type of tissue plasminogen activator variant. In this study, preplasmic and cytoplasmic (as inclusion body: IBs) production and activity of recombinant reteplase in E. coli were investigated and compared using a pET system (pET22b and pET15b). The cDNA of reteplase was cloned by polymerase chain reaction (PCR) amplification, sequenced, inserted into the vector pET 22b and pET15b, and expressed using isopropyl β-D-1-thiogalactopyranoside (IPTG). The recombinant plasmid was expressed in the form of inclusion body in pET 15b and in periplasmic space in pET22b. The obtained results of inclusion body extraction from recombinant pET22b (rpET22b) and recombinant pET15b (rpET15b) plasmids using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) showed a band of ∼39 kD. However, the obtained results of periplasmic space extraction from rpET22b plasmid showed a very weak band, while cytoplasmic expression of reteplase (pET15b) produced a strong protein band confirmed with Western blotting. Consequently, our results demonstrated that the cytoplasmic expression system is efficient for the production of reteplase protein in prokaryote systems and a high amount of reteplase was obtained from the expressed proteins in the form of IBs. The obtained activity of rpET15b plasmid showed a higher enzyme absorbance in comparison to rpET22b plasmid. This suggests rpET15b as an appropriate candidate for reteplase production.
Fatemeh Khodabakhsh; Mohammad Farid Zia; Fatemeh Moazen; Mohammad Rabbani; Hamid Mir Mohammad Sadeghi
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Preparative biochemistry & biotechnology     Volume:  43     ISSN:  1532-2297     ISO Abbreviation:  Prep. Biochem. Biotechnol.     Publication Date:  2013 Oct 
Date Detail:
Created Date:  2013-06-17     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  9607037     Medline TA:  Prep Biochem Biotechnol     Country:  England    
Other Details:
Languages:  eng     Pagination:  613-23     Citation Subset:  IM    
a Department of Pharmaceutical Biotechnology and Isfahan Pharmaceutical Sciences Research Center , School of Pharmacy and Pharmaceutical Sciences, Isfahan University of Medical Sciences , Isfahan , I.R , . Iran.
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