Expression And Antigenic Study Of A Fusion Protein Originated From Brucella Ribosomal Protein L7/L12and BLS

Brucellosis is a severe zoonosis caused by Brucella, a group of gram negativebacteria. The human symptoms include persistent fever, arthritis, andhepatosplenomegaly. Brucella infected animals suffer even worse effects such asabortion, infertility, and tissue necrosis. Coming to the21century, the infection andmorbidity rates stay high in some areas in our country and all over the world; thusBrucellosis has become one of the key factors that seriously threaten the health ofpeople and the economic development of affected regions. Currently, few thoroughcures have been reported around the world, and the most effective prevention isvaccination.The present study focused on a fusion protein originated from the Brucellaribosomal protein L7/L12and the Lumazine synthase (BLS) for the purpose toenhance the antigenecity. The two coding genes L7/L12and bls were fused by PCRamplified using fusion primers with genomic DNA of attenuated live vaccine S2of B.suis as template. The fusion gene was inserted into a expression vector resulted in arecombinant plasmid pET-28a-L7/L12-bls, and was expressed into a fusion proteinL7/L12-BLS with a molecular weight35.8kDa in engineered E. coli strain TL129under an IPTG inducible promoter control and the expressed protein was detected bySDS-PAGE. In addition, L7/L12gene was inserted into pET-28a resulted in arecombinant expression vector pET-28a-L7/L12that expressed a L7/L12protein withmolecular weight19.3kDa.Both L7/L12protein and L7/L12-BLS protein expressed in E. coli were purifiedby Ni-NTA agarose column. The purified proteins and PBS as black control wereused to inoculate long-eared rabbits by injection of diluted proteins. The antiserumswere collected and the immunogenecities were analysed by Western blot andEnzyme linked Immunosorbent Assay (Elisa). Results showed that the antiserumsagainst the two proteins had intense immune response to relevant proteins whileblack control had no any immune response. Furthermore, the antiserum againstL7/L12-BLS protein showed a two to five times stronger immune responsecompared to the antiserum against L7/L12, indicating that BLS protein had anenhancement effect.This study provided theoretical and experimental basis for the further development of gene vaccine and recombinant subunit vaccine.