Transduction of human Cu, Znª²superoxide dismutase mediated by an PEPª²1 peptide into endothelial cells of human umbilical vein

    DING Peng, WANG Jiaª²Ning, HUANG Yongª²Zhang, LUO Liª²Na, SHAO Fang

    Institute of Clinical Medicine, People¬ðs Hospital, Yunyang Medical College, Shiyan 442000, China, General Hospital of Pingdingshan Coalmine Group, Pingdingshan 467000, China

    ¡¾Abstract¡¿ AIM: To construct a prokaryotic expression system of pET15bª²PEPª²1ª²SOD1, and to determine whether the expressed PEPª²1ª²SOD1 fusion protein can be transduced into cells. METHODS: SOD1 cDNA was amplified by PCR using pBluescript II SKª²SOD1 as template, and digested with restriction endonuclease to construct recombinant plasmids pET15bª²SOD1 and pET15bª²PEPª²1ª²SOD1, followed by being respectively transformed into E.coli to produce SOD1 and PEPª²1ª²SOD1 fusion proteins, which were purified and added in cultured endothelial cells of human umbilical vein in vitro. RESULTS: The highly purified and active SOD1 and PEPª²1ª²SOD1 fusion proteins were obtained.The PEPª²1ª²SOD1 fusion protein could enter endothelial cells of human umbilical vein in a timeª² and doseª²dependent manner when added exogenously in a culture media. Once inside the cells, PEPª²1ª²SOD1 protein was enzymatically stable for 48 h. CONCLUSION: PEPª²1ª²SOD1 fusion protein can be transduced effectively into endothelial cells of human umbilical vein, which provides a basis for future study involving direct delivery of SOD1 protein into tissues for therapy. ......