A Hepatitis B Surface Antigen-binding Protein (sbp), Prokaryotic Secretory Expression, Purification And Mechanism Of The Initial Study

The current HBV vaccine performed certain protective role in preventing the infection of HBV. In our previous study, we have found a protein named HBsAg Binding Protein (SBP), which can specifically interact with HBsAg. Several experiments have proved SBP could enhance the immuno-response to HBV vaccine.In the present study, SBP gene was cloned into the prokaryotic expression vector pET22b and transformed into E.coli. strainBL21. The gene was inserted the downstream of IPTG inducible lacl promoter and pelB peptide signal sequence, so the protein was expressed into the periplasmic space. We also studied on several factors which play important parts on SBP expression level, and found that when SBP expression was induced 12 hours under IPTG concentration of 0.5mM, high expression level could be acquired. Subsequently SBP was purified through affinity chromatography and identified through Western Blot. Further experiment showed that SBP has the ability to bind specifically with HBsAg and calculated the affinity constant value.In the second part, a fusion protein, DsRed (red fluorescent protein) and SBP was constructed, which could trace SBP. The result showed that DsRed-SBP could specifically bind to HBsAg. Also, the luminous intensity and protein content showed a linear relationship. Flow cytometry and Western-Blot test confirmed SBP could bind receptors on cells, such as CD64/CD32/FcRn. Antibodies of CD64/CD 32/FcRn could block the SBP induced hepatitis B virus endocytosis.