关键词: 细胞穿透肽, 蛋白转导域, 蛋白转导, PEP-1, 增强型绿色荧光蛋白

Abstract: Objective To investigate the in vivo transduction capability of fusion protein PEP-1-EGFP with mice. 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. The expressed EGFP and PEP-1-EGFP were purified with Ni2+-resin affinity chromatography. Five hundred micrograms of EGFP and PEP-1-EGFP fusion protein were injected into mice from the caudal vein, respectively, the mice were euthanized and perfused with PBS after administration 2 hours later. Then, the heart, brain, liver, spleen and kidney were removed and sectioned with a cryostat at 7μm for visualization with a inverted fluorescent microscope. Results The brain, heart, liver, spleen and kidney in the mice injected with PEP-1-EGFP demonstrated bright and homogenous green fluorescence whereas that with EGFP showed no green fluorescence at all. Conclusions The successful expression and purification of PEP-1-EGFP fusion protein and its efficient transduction into mouse in vivo provide a basis for the research on transmembrane delivery of macromolecule drugs mediated by the cell-penetrating peptide, PEP-1, in molecular therapy of many diseases such as ischemia-reperfusion injury and tumor.

Key words: cell-penetrating peptide, protein transduction domain, protein transduction, PEP-1, enhanced green fluorescent protein