Over Expression of Influenza Virus M2 Protein in Prokaryotic System

Abstract

Background and Aims: Influenza A virus of Orthomyxoviridae family is able to create pandemic influenza. Vaccination is the most effective way to prevent influenza virus infection. Matrix protein 2 (M2) is a homotetramer ion channel with 97 amino acids length and highly conserved among influenza viruses and is considered for development of a universal influenza vaccine. Materials and Methods:We present here cloning and expression of influenza A virus M2 protein as a fusion with 6-His tag in Escherichia coli BL21 strain. The gene was amplified by PCR and ligated into the prokaryotic expression vector pET28a. The expression of M2 protein was induced by IPTG and confirmed by SDS-PAGE and western blotting. The desired protein was purified with affinity chromatography on a Ni-TED resin column and has to be evaluated in animal models for further studies. Results:The results of sequencing showed that M2 gene was cloned in pET28a properly in frame to histidine tag and the product was confirmed by xpreimmune reaction of monoclonal anti-M2 antibody to recombinant M2 in western blotting. Conclusion: This study might provide a basis for production of a universal and broadspectrum human influenza vaccine.

Enhancing the Yield and Quality of Influenza Virus-like Particles (VLPs) Produced in Insect Cells by Inhibiting Cytopathic Effects of Matrix Protein M2

“…Cellular expression of HA, M1, and M2 protein was quantified by Western blot analysis of cell lysates using purified HA (BEI Resources, Manassas, VA), M1 (Sino Biological, Beijing, China) and M2 (purified from E. coli as described) 74 as protein standards. Antibodies used include anti-HA (Sino Biological), anti-M1 (Invitrogen), and anti-M2 (Invitrogen) antibodies, alkaline phosphatase-conjugated antimouse or antirabbit IgG secondary antibody (Life Technologies), and antitubulin antibody (BioLegend).…”