Designing, Optimizing, and Structure Prediction of Chimeric Protein CTB-IpaD for Expression in E.coli

Seyed Mohammad Gheibi Hayat¹, Azam Sadeghinia^2*, Babak Elyasifar³, Ghasem Abbaszadeh-Goudarzi⁴, Zahra Adeli⁵ and Somaieh Salimi⁶

¹Young Researchers and Elite Club, Andimeshk Branch, Islamic Azad University, Andimeshk, Iran. ²Department of Cardiology, Faculty of Medicine, Dezful University of Medical Sciences, Dezful, Iran. ³Department of Drug Biotechnology, Faculty of Pharmacy, Tabriz University of Medical Sciences, Tabriz, Iran. ⁴Department of Medical Biotechnology, Faculty of Advanced Technologies in Medicine, Tehran University of medical Sciences, Tehran, Iran. ⁵Department of Microbiology, Islamic Azad University, Damghan Branch, Damghan, Iran ⁶Department of Marine Biology, Faculty of Marine Science, Chabahar Marine University, Chabahar, Iran.

DOI : http://dx.doi.org/10.13005/bbra/2253

ABSTRACT: The most common cause of diarrhea is Shigella and no vaccine has been found to provide protection against it. IpaD play an important role in invasion, infection, and pathogenesis caused by Shigella. This protein has been one of the most important protein for shigella vaccine candidate. This study aims to link the gene sequence IpaD and CTB with In-silico analysis. Nucleotide sequences were obtained from NCBI database and optimized. Sequences were fused together by a flexible linker in order to find the best epitope exposing chimeric antigen. After Protein half-life and instability index were determined, and then the prediction of the secondary structure and the three-dimensional structure was analyzed and also immunodominant linear B-cell epitopes were also settled at the final step. Codons were changed with respect to codon bias of E.coli and the GC content was changed to an optimal level. Inappropriate structures of RNA were removed. However, these changes led to a prolonged half-life and increased mRNA expression of the recombinant protein. Sequences of the chimeric gene was optimized based on codon usage pattern, In-silico analysis represented that sequence can have a high expression in E.coli and designed to enhance the expression of proteins which contain rare codons at high frequency. (EAAAK) 4 hydrophobic linkers could prevent the domain interactions of the chimeric protein. Chimeric protein can be a good candidate in immunogenicity study to produce an effective vaccine.

KEYWORDS: Shigella dysenteriae; IpaD protein; B subunit cholera toxin; Bioinformatics

This work is licensed under a Creative Commons Attribution 4.0 International License.