细胞穿透肽PEP-1介导增强型绿色荧光蛋白转导人结直肠癌SW480细胞

背景与目的:细胞穿透肽是近年来发现的一类能介导大分子物质跨膜转导的小分子肽段,本实验研究细胞穿透肽PEP-1携带大分子蛋白穿透人结直肠癌细胞SW480的能力.方法:用基因工程的方法制备并纯化EGFP蛋白和PEP-1-EGFP融合蛋白,将纯化后的两种蛋白与体外培养的人结直肠癌细胞共同孵育,在荧光显微镜下直接观察PEP-1介导目的蛋白EGFP转导入SW480细胞的能力.结果:EGFP蛋白不能进入细胞内,PEP-1-EGFP融合蛋白和SW480细胞共同孵育1h后可见有明亮绿色荧光分布于胞浆和胞核.结论:PEP-1能有效携带目的蛋白进入人结直肠癌细胞.

Cell-penetrating peptide PEP-1-mediated transduction of enhanced green fluorescent protein into human colorectal cancer SW480 cells

BACKGROUND & OBJECTIVE: Cell-penetrating peptides, a class of small cationic peptides, could mediate macromolecules transduction into many cell lines. This study was to explore the penetrating ability of PEP-1 in the transduction of enhanced green fluorescent protein (EGFP) into human colorectal cancer cell line SW480. METHODS: Two prokaryotic expression plasmids pET15b-EGFP and pET15b-PEP-1-EGFP were constructed and transformed into E.coli BL21 (DE3) to express EGFP and fusion protein PEP-1-EGFP, respectively. Purified EGFP and PEP-1-EGFP were incubated with SW480 cells. The expression of EGFP in SW480 cells was observed under inverted fluorescent microscope. RESULTS: EGFP could not be transduced into SW480 cells, whereas PEP-1-EGFP fusion protein could be transduced into SW480 cells and distributed in cytoplasm and nuclei after 1-hour incubation. CONCLUSION: Mediated by PEP-1, EGFP could be transduced efficiently into SW480 cells.