title: Fused hydrophobic elastin-like-peptides (ELP) enhance biological activity of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL).
abstract: Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising anticancer agent with tumor-selective apoptotic activity. Formation of aggregates as trimer is the prerequisite for TRAIL's function as an apoptosis inducer. However the polymerization property of TRAIL has also brought difficulties for its production. RGD-TRAIL is an integrin-targeting TRAIL mutant with enhanced apoptosisinducing activity towards tumor cells both in vitro and in vivo. When expressed in E. coli, TRAIL or its mutant RGDTRAIL usually formed inclusion bodies. Their extreme aggregation propensity for aggregation destabilizes the protein, leading to poor recovery and therefore low yield from the purification process. The low purification efficiency of TRAIL retards its industrial application and large-scale production. To avoid the above problems during RGD-TRAIL production, we employed elastin-like polypeptides (ELPs) for the fusion-expression of recombinant RGD-TRAIL. Recombinant RGD-TRAIL-ELP was expressed in a soluble form and efficiently purified from the clarified cell extracts by three rounds of inverse transition cycling (ITC). SDS-PAGE and Western blotting analyses of purified RGD-TRAIL-ELP showed that RGD-TRAIL-ELP was successfully purified and the yield was up to 10 mg/L of bacterial culture. Apoptosis assay was performed in human colorectal carcinoma cells (COLO-205) and human breast cancer cell line (MDA-MB-231) to assess the potency of the fusion protein. Fusion with hydrophobic ELP effectively enhanced RGD-TRAIL's biological activity. The higher activity and appropriate particle size of RGD-TRAIL-ELP could be used for RGD-TRAIL delivery in tumor therapy.
authors: Kaizong
authors: K
authors: Huang
authors: Ningjun
authors: N
authors: Duan
authors: Wenyan
authors: W
authors: Zou
authors: Chunmei
authors: C
authors: Zhang
authors: Yueyang
authors: Y
authors: Lai
authors: Pingping
authors: P
authors: Shen
authors: School of Life Sciences, Nanjing University, 163 Xianlindadao Road, Nanjing 210023, Jiangsu, People's Republic of China. zchua@nju.edu.cn.
authors: Zichun
authors: Z
authors: Hua
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D045744:Cell Line, Tumor; D004549:Elastin / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000737:chemistry / Q000378:metabolism*
chemical_list: D010455:Peptides; D011993:Recombinant Fusion Proteins; D053221:TNF-Related Apoptosis-Inducing Ligand; D004549:Elastin