[Cloning, soluble expression and characterization of human sBCMA].
BCMA is one of the transmembrane receptors belonging to BAFF and APRIL. In order to identify the feasibility of sBCMA as decoy receptor and obtain active sBCMA for its structural and functional research, full length of hBCMA was amplified with total RNA from Raji cell line by RT-PCR, and the cDNA encoding the extracelluar soluble domain of hBCMA was inserted into pET43.1a(+) vector. The recombinant vector pET43.1a(+)-sBCMA was transformed into E. coli Origami B(DE3) pLyS which is helpful for disulfide bond construction of expression proteins. After IPTG induction, the recombinant protein was expressed as soluble fusion protein, sBCMA-NusA-His6, and identified by western blotting. Then the target protein was purified by Ni(+)-chelating Sepharose Fast Flow. The binding activity between recombinant sBCMA and BAFF was detected by ELISA. Also, Recombinant sBCMA inhibited proliferation of mouse B cell stimulating by rhsBAFF. It was proved that recombinant sBCMA has good bioactivity and the method to express those proteins rich in disulfide bond is feasible and effectual.
Guan
Zheng-Bing
ZB
Jiangsu Province Key Laboratoryfor Molecular and Medical Biotechnology, Life Sciences College, Nanjing Normal University, Nanjing 210097, China.
Cao
Peng
P
Ye
Ji-Lin
JL
Zhang
Shuang-Quan
SQ
D053264:B-Cell Activating Factor / Q000737:chemistry; D053301:B-Cell Maturation Antigen / Q000096:biosynthesis / Q000235:genetics*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D004220:Disulfides / Q000737:chemistry; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006801:Humans; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics; D012995:Solubility; D053300:Tumor Necrosis Factor Ligand Superfamily Member 13 / Q000737:chemistry
D053264:B-Cell Activating Factor; D053301:B-Cell Maturation Antigen; D018076:DNA, Complementary; D004220:Disulfides; D011993:Recombinant Fusion Proteins; D053300:Tumor Necrosis Factor Ligand Superfamily Member 13