Preliminary Research On The Anti-tumor Activities And Transduction Of Different Cell Penetrating Peptides

While a variety of transporters currently exist for cargo translocation, cell-penetrating peptides (CPPs) are typically short cationic sequences, which have become one of the most popular and efficient techniques for achieving intracellular access. CPPs were labeled with apoptin (VP3) and enhanced green fluorescent protein (EGFP). These different CPP fusion proteins were obtained after appropriate expression and purification. Then CPP fusion proteins HBD and TAT were connected with VP3 for comparative pharmacological experiment, while CPPs labeled with EGFP were traced in different cell lines to make a comparative study on the efficiency and specificity of transduction activity. HBD as a new CPP had certain transduction activity by these two studies.HBD-VP3-pET28a and TAT-VP3-pET28a, which were expression vectors of CPP fusion proteins, had been successfully constructed by our lab.Following induction of expression, HBD-VP3 and TAT-VP3 were efficiently purified by denaturation and renaturation of the inclusion body. Then the inhibit experiment in vitro (MTT assay) was conducted to observe the growth of tumor cells in two groups. The results indicated that both HBD and TAT could obviously induce apoptosis of HeLa cells. However, transduction efficiency of HBD showed relatively lower than TAT in delivering VP3 into HeLa cell.In this paper, EGFP as reports gene was coupled to sequence of different CPPs (HBD or R7) respectively. Cloned HBD-EGFP and R7-EGFP fusion protein were expressed and purified by Ni2+-NTA affinity chromatography. Transduction activity of HBD and R7 was observed and compared in different tumor cells. In the study presented here, we have explored the efficiency and specificity of coupling EGFP to R7 and HBD crossing the membrane of five cell lines. The study indicates that R7 and HBD are good peptide vectors to deliver EGFP into these five cells with different incubation time. However, compared with R7, the efficiency of HBD for carrying EGFP into tumor cells was relatively lower. Further considerations are needed to improve the efficiency of transmembrane protein HBD,and thus effective HBD mutants are being designed.