Generation of a novel TRAIL mutant by proline to arginine substitution based on codon bias and its antitumor effects.

Generation of a novel TRAIL mutant by proline to arginine substitution based on codon bias and its antitumor effects. Mol Med Rep. 2017 Aug 02;: Authors: Zhu A, Wang X, Huang M, Chen C, Yan J, Xu Q, Wei L, Huang X, Zhu H, Yi C Abstract TNF ligand superfamily member 10 (TRAIL) is a member of the tumor necrosis factor superfamily. The present study was performed in an effort to increase the expression of soluble (s)TRAIL by rebuilding the gene sequence of TRAIL. Three principles based on the codon bias of Escherichia coli were put forward to design the rebuild strategy. Relying on these three principles, a P7R mutation near the N‑terminal region of sTRAIL, named TRAIL‑Mu, was designed. TRAIL‑Mu was subsequently cloned into the PTWIN1 plasmid and expressed in E. coli BL21 (DE3). Using a high‑level expression system and a three‑step purification method, soluble TRAIL‑Mu protein reached ~90% of total cellular protein and purity was >95%, demonstrating success in overcoming inclusion body formation. The cytotoxic effect of TRAIL‑Mu was evaluated by sulforhodamine B assay in the MD‑MB‑231, A549, NCI‑H460 and L02 cell lines. The results demonstrated that TRAIL‑Mu exerted stronger antitumor effects on TRAIL‑sensitive tumor cell lines, and was able to partially reverse the resistance of a TRAIL‑resistant tumor cell line. In addition, TRAIL‑Mu exhibited no notable biological effects in a normal liver cell line. ...
Source: Molecular Medicine Reports - Category: Molecular Biology Tags: Mol Med Rep Source Type: research