Expression, purification and identification of recombinant human lymphotoxin α deletant (rhLT-αΔN27)

AIM: To construct prokaryotic expression vector of recombinant human lymphotoxin α deletant (rhLT-αΔN27) and express the protein in E.coli. METHODS: The rhLT-αΔN27 gene was amplified by RT-PCR using total RNA extracted from Jurkat cells, cloned into prokaryotic expression vector pET-23b, and transformed into E.coli BL21(DE3). The recombinant protein was expressed after IPTG induction and purified by DEAE Sepharose FF and Phenyl-Sepharose FF. RESULTS: The recombinant protein was expressed as inclusion bodies with the yield of more than 30% of total bacterial protein. After purification, the purity of rhLT-αΔN27 was 99%, and the biological activity was more than 8×10 7 U/mg. Other characteristics of rhLT-αΔN27, such as relative molecular mass(M r), pI and N-terminal amino acid sequence, all corresponded to theoretical prediction. CONCLUSION: The expression vector of rhLT-αΔN27 gene was constructed, and the recombinant protein was expressed in E.coli successfully. A method of for purifying rhLT-αΔN27 was established.