Fig. 1. Development of ILz(6):TRAIL. (A) Schematic representation of the ILz(6):TRAIL protein. The 6xHis-tag was essential for Ni-NTA affinity purification; ILz(6) represents the isoleucine zipper hexamerization motif and TRAIL (114-281) represents the extracellular region of human TRAIL amino acids 114-281; (B) Purification of the recombinant ILz(6):TRAIL protein. The bacterial expression vector pET23a(+):ILz(6): TRAIL was transformed into E. coli BL21 (DE3) cells and induced with IPTG. Recombinant ILz(6):TRAIL was purified using a Ni-NTA His-binding affinity column; (C) HCT116 cells were treated overnight with recombinant ILz(6):TRAIL following by recording light microscope images; (D) Activity analysis of recombinant ILz(6):TRAIL. BJAB cells were treated with the the indicated amounts of purified recombinant ILz(6):TRAIL for 2 hours and cell death was determined using the XTT assay; (E) Comparison of the apoptosis-inducing activities of TRAIL and ILz(6):TRAIL. BJAB cells were treated with recombinant TRAIL or ILz(6): TRAIL for 18 hours and cell death was measured using the XTT assay.