Construction of Prokaryotic Expression Vector for Ag85A-HA2 Fusion Gene and Studies on the Immunity Efficacy of Fusion Protein Against Influenza a Virus

SHAO Jing-jing, YANG Jing, DAI Jun. et al

Abstract

To construct Ag85A-HA2 prokaryotic expression vector, express the fusion protein and study the immunity efficacy of fusion protein against influenza A virus. Methods Ag85A-HA2 prokaryotic expression vector was constructed and induced with IPTG．The fusion protein was identified by SDS-PAGE and purified with His-Tag affinity chromatography. The BALB/c mice were immunized with fusion protein. Then the pathological section, lung index, lung inhibitory rate and death-protection rate were tested to evaluate the immunity efficacy of fusion protein. Results pET-32a(+)/Ag85A-HA2 prokaryotic expression vector was constructed successfully. And SDS-PAGE indicated that fusion protein was expressed correctly with a molecular mass of 70×103. The lung index and death-protection rate in experimental group were 39.30% and 80%, higher than that of control group. The pathological section also demonstrated that Ag85A-HA2 fusion protein had a protective effect on murine lungs. Conclusion Ag85A-HA2 prokaryotic expression vector was successfully constructed, inducible expression and the fusion protein had an immunity efficacy against influenza A virus in animal experiment.

 

Keywords:  Influenza, A virus, Ag85A Hemagglutinin (HA), Prokaryotic expression Immunity efficacy 

 

Full Text:

PDF


References


Steel J, Lowen AC, Wang TT, et al. Influenza virus vaccine based on the conserved hemagglutinin stalk domain. MBio, 2010; 1( 1):pii;e00018-10. doi: 10. 1128/mBio. 0018-10.

Bommakanti G, Citron MP, Hepler RW, et al. Design of an HA2-based Escherichia cov expressed influenza immunogen that protects mice from pathogenic challenge. Proc Natl Acad Sci USA,2010; 107(31): 13701-13706.

Sandbulte MR, Westgeest KB, Gao J, et al. Discordant antigenic drift of neuraminidase and hemagglutinin in H1N1 and H3N2 influenza viruses. Proc Natl Acad Sci USA, 2011; 108 (51):20748-20753.

Romano M, Roupie V, Hamard M, et al. Evaluation of the immunogenicity of pBudCEl. 1 plasmids encoding mycolyl- transferase Ag85A and phosphate transport receptor PstS-3 from Mycobacterium tuberculosis. Vaccine, 2006; 24(21): 4640-4643.

Dai J, Pei D. Wang B, et al. A novel DNA vaccine expressing the Ag85A-HA2 fusion protein provides protection against influenza A virus and Staphylococcus aureus. Virol J ,2013; 10 : 40-52. doi: 10. 1186/1743-422X-10-40.

Dessing MC, van der Sluijs KF, Florquin S, et al. Toll-like receptor 2 does not contribute to host response during postinfluenza pneumococcal pneumonia. Am J Respir Cell Mol Biol, 2007; 36(5): 609-614.

van der Sluijs KF, van Elden LJ, Nijhuis M, et al. IL-10 is an important mediator of the enhanced susceptibility to pneumococcal pneumonia after influenza infection. J Immunol. 2004;172(12);7603-7609.

MiKi K, Nagata T, Tanaka T, et al. Induction of protective cellular immunity against Mycobacterium tuberculosis by recombinant attenuated self-destructing Listeria monocytogenes strains harboring eukaryotic expression plasmids for antigen 85 complex and MPB/MPT51 . Infect Immun,2004;72(4) :2014- 2021.

Forg P. von Hoegen P, Dalemans W. et al - Superiority of the ear pinna over muscle tissue as site for DNA vaccination. Gene Ther, 1998;5(6) :789-797.

Ayash-Rashkovsky M. Weisman Z.Zlotnikov S,et al. Induction of antigen-specific Thl-biased immune responses by plasmid DNA in schistosoma-infected mice with a preexistent dominant Th2 immune profile. Biochem Biophys Res Cornmun.2001;282 (5) ; 1169-1176.

Ynan EF, Webster RQ, Fuller DH, et al. DNA vaccines; protective immunizations by parenteral mucosal, and gene-gun inoculations. Proc Natl Acad Sci USA, 1993; 90 ( 24 ); 11478- 11482.

McMahon JM. Wells DJ. Electroporation for gene transfer to skeletal muscles:current status. Bio Drugs. 2004; 18(3): 155- 165.


Refbacks

  • There are currently no refbacks.