Objective: To construct a prokaryotic expression vector of the His-tagged truncated factor H-binding protein (Fhb) fragments, Fhb-N (amino acids 45-344aa) and Fhb-C (amino acids 345-644aa), of Streptococcus suis serotype 2, express it in E.coli BL21 (DE3) in order to acquire high-purity recombinant protein, and finally identify the binding activity with human serum IgG (hIgG).
Methods: Fhb-N gene and Fhb-C gene were amplified using the primers designed according to 05ZYH33 genome sequences and cloned into the expression vector pET28a⁺ to construct recombinant plasmids. The plasmids were transformed into E.coli BL21 (DE3) and induced to express by IPTG. The recombinant proteins were purified by nickel affinity chromatography and identified by Western blotting. The hIgG was purified from human serum by HiTrap protein G HP column in accordance with the manufacturer's instructions. In addition, the specific binding to hIgG was identified by Western blotting and biolayer Interferometry (BLI).
Results: The prokaryotic expression vector of His-Fhb-N and His-Fhb-C was constructed, and the target proteins were expressed, purified and identified. The specific binding activity with hIgG was identified and the binding region was found located on the Fhb-N(45-344aa).
Conclusion: His-Fhb-N can specifically bind to hIgG, which will help us to further study the role of Fhb-hIgG interaction in the pathogenesis of Streptococcus suis.