High copy numbers and N terminal insertion position of influenza A M2E fused with hepatitis B core antigen enhanced immunogenicity.
The extra domain of influenza M2 protein (M2e) is almost completely conserved among all influenza A virus subtypes. M2e is a promising candidate target for the development of a broad-spectrum recombinant influenza A vaccine. However, the immunogenicity of M2e needs to be improved. Copy numbers of M2e and its fusion expression with different carrier proteins may affect its immunopotency. In this study, we designed and created different constructs through genetic fusion of M2e (MSLLTEVETPTRSEWECRCSDSSD) (A/California/05/2009 (H1N1)) with the N-terminus (HBcAg1-149aa + Cys) by insertion in the N-terminus Hepatitis B Core (HBc) antigen 1-149aa and Middle 78-81aa of HBcAg1-149aa to construct a recombinant M2e-based vaccine candidate. These chimeric sequences were expressed in Escherichia coli. We constructed fusion proteins containing influenza A H1N1 influenza virus (2009), as well as one, two, and three copies of M2e and hepatitis B core antigen1-149aa amino acid-optimized codon inserted N and its intermediate. The recombinant protein was expressed and purified. Western blot analysis was employed to evaluate the expression of the M2e recombinant protein containing different copy numbers of M2e. Mice were immunized for two times with the purified fusion protein HBc/M2e BALB/c. Serum levels of M2e antibody gradually increased along with increase in immunity. The levels of different fusion protein M2e antibodies increase with increasing M2e copy number. In addition, the protein antibody level in the N terminal fusion protein is higher than that in intermediate fusion.
Basic Medical School of Zhengzhou University.
Xincheng
X
Sun
Yunlong
Y
Wang
Caiwen
C
Dong
Jinqiang
J
Hu
Liping
L
Yang
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D007252:Influenza Vaccines / Q000737:chemistry* / Q000276:immunology*; D051379:Mice; D008969:Molecular Sequence Data; D010957:Plasmids / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification; D015203:Reproducibility of Results; D044967:Serum / Q000378:metabolism
D007252:Influenza Vaccines; C509609:M2e-HBc influenza vaccine; D011993:Recombinant Fusion Proteins