| Objective:To clone and express heat-shock protein40gene of Schistosoma japonicum (SjHSP40) and analyze its immunological characteristics.Methods:First, the gene encoding SjHSP40was analyzed by bioinformatic methods and this fragment of DNA was amplified by PCR. The gene was subcloned into the prokaryotic expression vector, pGEX-6P-1. The recombinant plasmid pGEX-6P-1-SjHSP40was transformed into E. coli BL-21(DE3) and the protein expression was induced with IPTG. Then, the recombinant protein was purified with Glutathione-Sepharose4B resin and analyzed by SDS-PAGE and Western-blot. Finally, we immunized the mice, Balb/c strain, with the fusion protein. Following that, the titer of specific antibody in different isotypes, including IgGl and IgG2a, in mouse serum was determined by ELISA.The frequencies of Thl、Th2、Th17and Treg cell populations in total CD4+T cells were analyzed by flow cytometry.Results:Bioinformatic analyzsis showed that the closest homologue with SjHSP40was the HSP40from S. mansoni. The amino acid sequence was86%,37%and39%identified to its homologue protein in S. mansoni, in Homo sapience and in Mus musculus, respectively. Seven B cell epitopes and many T cell epitopes were predicted with BepiPredl.O sever and IEDB sever, respectively. DNA sequencing showed that the recombinant plasmid, pGEX-6P-1-SjHSP40, was successfully constructed. The fusion protein of GST-SjHSP40was produced, purified and specifically recognized by anti-GST antibody. Compared to GST control groups, we found that this fusion protein significantly induced the production of specific antibody in different isotypes, including IgG1and IgG2a, in mouse serum. However, the results showed that SjHSP40had no significant influence on the frequencies of Th1、Th2、Th17and Treg cells in total CD4+T cells.Conclusions:SjHSP40significantly up-regulated the production of specific antibody in the serum of the immunized mice, while no significant change appeared on CD4+T cell. |
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