Preparation and functional analysis of recombinant human BAFF_(112-285)

Objective To lay a foundation for further study on B cell activating factor belonging to the TNF family (BAFF) by prepa ring the fun ctiona l recombinant extracellular region 112-285 amino acids of human BAFF (rhBAFF 112-285 ). Methods Total RNA was extracted from human HL 60 ce lls and the cDNA encoding human BAFF extracellular region 112-285 amino acids (BAFF 112-285 ) was amplified by RT PCR. After sequenced, the cDNA was cloned into the prok aryotic expression vector pQE 80L and the recombinant vector was transformed in to E.coli .DH5α. The rhBAFF 112-285 was expressed by the transformed c ells induced by IPTG, and was purified with Ni 2+ NTA chromatography. Then the immunological activity of the purified protein was assayed by using 3 H TdR i ncorporation test. Results A 525 bp cDNA was amplified by RT PCR, which was consistent with the sequence encoding human BAFF 112-285 am ino acids reported in GenBank. The rhBAFF 112-285 was highly expressed by E.c oli , which could reach 45.7% of total bacterial proteins. The purity of the pu rified protein could reach 98.4%. The purified rhBAFF 112-285 could effici ently activate human peripheral lymphocyte. Conclusion The rhBA FF 112-285 can be highly expressed by E.coli . The purified protein pos sesses biological activity, which may pave a way for further studies.