Fusion and sequence analysis of the influenza A (H۹N۲) virus M۲e and C-terminal fragment of Mycobacterium tuberculosis HSP۷۰ (H۳۷Rv)

The present study was aimed to construct a fusion plasmid harboring the extracellular domain of the influenza A M۲-protein (M۲e), which was fused to the N-terminus of the truncated HSP۷۰ (HSP۷۰۳۵۹–۶۱۰) molecule as a new approach for future vaccine research against influenza A. The amplified fragments, M۲e and HSP۷۰۳۵۹-۶۱۰ genes, were gel-purified. The products were then single digested with BamHI restriction enzyme separately. The digested products were again gel-purified and ligated by T۴ DNA ligase to form M۲e- HSP۳۵۹-۶۱۰ gene. The PCR product containing both M۲e and HSP۳۵۹-۶۱۰ genes as a single open reading frame (ORF) was gel-purified and double digested with EcoRI and XbaI restriction enzymes and then ligated into the EcoRI / XbaI double digested pPICZαA expression vector to form recombinant expression vector. Finally, the fused gene was sequenced, and then confirmed according to the related deposited gene in Genbank. The extracellular domain of the M۲ protein, M۲e, which consists of N-terminal ۲۴ residues, showed to be remarkably conserved, and the N-terminal epitope SLLTEVET (residues ۲-۹) was conserved among all subtypes of influenza A viruses. Because of M۲e limited potency hence, low immunogenicity, it seems by linking this M۲e-peptide to an appropriate carrier such as mycobacterium tuberculosis C-terminal ۲۸-kDa domain of HSP۷۰ (hsp۷۰ ۳۵۹–۶۱۰) we can render it very immunogenic, but further study needs to express it in both prokaryotic and eukaryotic systems and then evaluate this fusion protein in animal model.

کلیدواژه ها:

Avian Influenza ، fusion ، vaccine ، HSP۷۰ ، M۲e