Expression And Purification Of Tp0453 Outer Membrane Recombinant Protein Of Treponema Pallidum And Characterization Of Its Immunocompetence

Objective:To construct a recombinant vector containting the gene encoding immuno-dominant epitope of Tp0453 outer membrane protein of Treponema pallidum (T. pallidum), and to express and purify Tp0453 outer membrane recombinant protein and analyze the immunoreactivity and immunogenicity of recombinant protein., to find a new antigen for the exploiting diagnosis of Tpallidum.Methods:The immuno-dominant epitope of Tp0453 gene was chosen by computer analysis , then was amplified from T. pallidum complete genome by polymerase chain reactions (PCR), subcloned into the expression vector pQE32 to generate recombinant plasmid pQE32/Tp0453, then expressed in E. coli M15, and analyzed by SDS-PAGE and Western-Blot. The fusion protein was purified with Ni-NTA affinity chromatography, Purified protein was analyzed by SDS-PAGE, and protein concentration was determined by the BCA protein assay kit. The immunoreactivity of fusion protein was evaluated by Indirect ELISA, Serum T. pallidum-anlibody was detected with indirected-ELISA based on the fusion protein. Zealand rabbits were immunized with the fusion protein, antibodies to anti-Tp0453 in sera were detected with indirected ELISA.Results:The size of PCR amplification product was about 800bp. Restriction enzyme digestion analysis and sequencing showed that the inserted target gene were Tp0453, compared with gene reported by GenBank, it had 100% similarity; and the results of BLAST confirmed that the sequence was Tp0453. A fusion protein with molecular...