Iranian Biomedical Journal
مجله بیومدیکال ایران
IBJ
Basic Sciences
http://ibj.pasteur.ac.ir
1
admin
1028-852X
2008-823X
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10.61882/ibj
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8888
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en
jalali
1400
8
1
gregorian
2021
11
1
25
6
online
1
fulltext
en
Production of an Antibody Fragment (scFv) Targeting PcrV Protein of Pseudomonas aeruginosa in Fed-Batch Cultivation Mode
Medical Biotechnology
Medical Biotechnology
مقاله کامل
Full Length/Original Article
<strong>Background: </strong><em>Pseudomonas aeruginosa</em> is one of the opportunistic pathogens causing frequent hospital-acquired life-threatening infections in mechanically ventilated patients. The most significant virulence factor of <em>P. aeruginosa</em> is type III secretion system (T3SS). PcrV is an important structural protein of the T3SS. <strong>Methods:</strong> In the current investigation, a recombinant single-chain fragment variable (scFv) mAb against the PcrV protein was expressed in EnBase® (fed-batch) cultivation mode. The pETiteTM N-His SUMO Kan vector, including anti-PcrV scFv gene, was transformed into <em>Escherichia coli</em> (BL21) cells. The expression and solubility of <a name="_Hlk50975731">anti-PcrV scFv </a>protein were investigated <a name="_Hlk50975698">at two different temperatures (25 </a>°C and 30 °C) and at different induction times (4, 6, 8, 12, and 24 hours). <strong>Results:</strong> Increased efficiency was achieved by EnBase® compared to Luria–Bertani broth; owing to the slow release of glucose, the maximum level of solubility and total protein expression was observed in EnBase® cultivation system at 30 °C and 24 h post induction. Furthermore, IC<sub>50</sub> for anti-PcrV scFv protein was determined to be approximately 7 μg/mL. <strong>Conclusion:</strong> Anti-PcrV scFv produced in this study showed promising <em>in vitro</em> results, protecting RBC from lysis by<em> P. aeruginosa</em> (<em>exoU+</em>).
Fed Batch, recombinant protein, Pseudomonas aeruginosa, scFv
390
398
http://ibj.pasteur.ac.ir/browse.php?a_code=A-10-3524-2&slc_lang=en&sid=1
Saba
Karam