Abstract:

 Objective   To prepare highly purified human soluble mutant B cell activating factor (smBAFF) and study its biological activity. Methods   Expression of the recombinant human smBAFF in E.coli BL21 was induced by IPTG, and the protein was assayed by SDS-PAGE and Western blot. The bacteria were split by sonication, inclusion bodies were extracted and dissolved, and then the smBAFF was purified by Ni2+-NTA affinity chromatography. The purified protein was refolded under specified conditions, and the bioactivity of the protein was assayed by immunofluorescence and CCK-8. Results   SDS-PAGE electrophoreses displayed that 1.7×104  recombinant protein was expressed, and Western blot showed that it was the protein of interest. The highly purified protein of the recombinant human smBAFF was attained by Ni2+-NTA affinity chromatography, and the recombinant protein could bind B cells, but lost the activity of co-stimulating B cell proliferation. Also, it could competitively inhibit the function of the soluble BAFF. Conclusions   The recombinant smBAFF was successfully prepared and possesses B cell binding activity but lost the activity of co-stimulating B cell proliferation. The result may lay a foundation to study the therapy for B cell malignancies and B cell abnormal activity diseases.

Key words: B cell activating factor belonging to the TNF family; Prokaryotic cells; Inclusion Bodies; Protein renaturation