title: Construction and expression of sTRAIL-melittin combining enhanced anticancer activity with antibacterial activity in Escherichia coli.
abstract: Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), as an anticancer protein with tumor-selective apoptotic activity, has been examined for use in clinical application. Melittin, an antibacterial peptide isolated from the bee Apis mellifera, has shown strong cytotoxicity to both tumor and normal cells. To ameliorate the cytotoxicity of melittin on cells and enhance the activity of TRAIL on cancer cells, we constructed a novel fusion protein, sTRAIL-melittin, containing a small ubiquitin-related modifier (SUMO) tag and expressed this fusion protein in Escherichia coli. Data showed that expression of the soluble fusion protein with the SUMO tag was approximately 85% of total target protein which was much higher than that without the SUMO tag (approximately 10%); sTRAIL-melittin was easily purified using Ni-NTA affinity chromatography and the tag was removed easily using SUMO-specific protease. To assay anticancer activity and side effects, methyl thiazolyl tetrazolium, hemolytic, and apoptosis assays were employed. Results demonstrated that sTRAIL-melittin had cytotoxic and apoptotic activity in K562 leukemia cells and HepG2 liver carcinoma cells, while it had only a minimal effect on erythrocytes and normal HEK293 cells. This indicates that the cytotoxicity of sTRAIL-melittin in normal cells was low and the anticancer activity of the fusion protein in tumor cells was significantly enhanced compared with sTRAIL (P<0.01). Furthermore, we found that sTRAIL-melittin also showed antibacterial activity to Staphylococcus aureus due to the presence of the melittin domain. Therefore, TRAIL fused with an antibacterial peptide may be a promising novel TRAIL-based anticancer treatment strategy.
authors.affiliation: Jiangsu Province Key Laboratory for Molecular and Medical Biotechnology, Life Sciences College, Nanjing Normal University, Nanjing, 210046, China.
authors.forename: Hongyan
authors.initials: H
authors.lastname: Liu
authors.forename: Yangyang
authors.initials: Y
authors.lastname: Han
authors.forename: Haiyan
authors.initials: H
authors.lastname: Fu
authors.forename: Meng
authors.initials: M
authors.lastname: Liu
authors.forename: Jing
authors.initials: J
authors.lastname: Wu
authors.forename: Xiaonan
authors.initials: X
authors.lastname: Chen
authors.forename: Shuangquan
authors.initials: S
authors.lastname: Zhang
authors.forename: Yuqing
authors.initials: Y
authors.lastname: Chen
mesh_terms: D000900:Anti-Bacterial Agents / Q000378:metabolism*; D000970:Antineoplastic Agents / Q000378:metabolism*; D017209:Apoptosis; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006801:Humans; D008555:Melitten / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013211:Staphylococcus aureus / Q000187:drug effects*; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000235:genetics / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D000970:Antineoplastic Agents; D011993:Recombinant Fusion Proteins; D053221:TNF-Related Apoptosis-Inducing Ligand; D008555:Melitten