中国生物工程杂志

Abstract: Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL, Apo-2L) is a mediator of cell death that preferentially targets cancer cells. The C-terminal extracellular region of Apo-2L (amino acids 114-281) exhibits a homotrimeric subunit structure. Soluble Apo-2L(sApo-2L) induces extensive apoptosis in lymphoid as well as non-lymphoid tumor cell lines. But it is limited in its efficacy by the short circulating half-life and low stability. sApo2L-Fc is a fusion protein consisting of Apo-2L and a fragment(hinge, CH2, CH3 domains) of the Fc of the IgG1. The DNA was constructed in the pcDNA3.1（+） eukaryotic expression vector. Using DOTAP-mediated gene transfer technique, pcDNA3.1- sApo2L-Fc was transformed into Chinese hamster ovary(CHO) cells. The selective medium containing G418 was then employed to select the positive colony. After purified with protein A affinity chromatography, the product was detected with SDS-PAGE and Western blot. A protein band with molecular weight of 44KDa was found in the SDS-PAGE. Western blot displayed the recombinant product had strong immunological activity with mouse anti-human Fc monoclonal antibody. After purification, the biological activity of sApo2L-Fc was more than 1.0×105IU/mg.