In-silico design and production of a novel antigenic chimeric Shigella IpaB fused to C-terminal of Clostridium perfringens enterotoxin
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The emergence of antibiotic-resistant phenotypes in Shigella serotypes and the high mortality rate, approximately one million dead annually, in affected patients announce a global demand for an effective serotype-independent vaccine against Shigella. This study aims to design, express, and purify a novel chimeric protein, as a serotype-independent vaccine candidate against Shigella containing full-length Shigella invasion plasmid antigen B (IpaB) and a C-terminal fragment (residues 194–319) of Clostridium perfringens enterotoxin (C-CPE) as a mucosal adjuvant. Several online databases and bioinformatics software were utilized to design the chimeric protein and the relative recombinant gene. The recombinant gene encoding IpaB–CPE194–319 was synthesized, cloned into pACYCDuet-1 expression vector, and transferred to E. coli Bl21 (DE3) cells. IpaB–CPE194–319 was then expressed in auto-induction medium, purified and characterized using MALDI-TOF-TOF mass spectrometry. Followed by subcutaneous injection of the purified IpaB–CPE194–319 to BALB/c mice, antigenicity of this chimeric protein was determined through performing dot-blot immunoassay on nitrocellulose membrane using mice sera. The outcomes of this study show the successful design, efficient expression, and purification of IpaB–CPE194–319 divalent chimeric protein under mentioned conditions. The obtained results also demonstrate the intrinsic antigenic property of IpaB–CPE194–319.
KeywordsShigella Vaccine candidate IpaB Chimeric protein Clostridium perfringens Enterotoxin CPE In silico
The authors would like to thank Dr. Iradj Ashrafi Tamai and Mr. Rasoul Sorbi for their cooperation during the project. We also tank Mrs. Bahareh Arabshahi for her helpful assistance in drawing and generating graphic works.
This study was funded by Shiraz University (Grant Number 9160423).
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
This study was approved by Animal Ethics Committee (AECs) of School of Veterinary Medicine, Shiraz University (permit: 94INT1M163973), and all the animal experiments were performed in accordance with guidelines and regulations of this committee (dated 20 September 2013) adhering to the Helsinki Declaration.
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