A gene delivery system based on the N-terminal domain of human topoisomerase I

作者：Chen Yi An; Kuo Hsiao Che; Chen Young Mao; Huang Shin Yi; Liu Yu Ru; Lin Su Ching; Yang Huey Lang; Chen Tzong Yueh^*

来源：Biomaterials, 2011, 32(17): 4174-4184.

DOI：10.1016/j.biomaterials.2011.02.041

• Gene delivery system
• Protein transduction domain
• Topoisomerase I
• Membrane penetration domain
• DNA binding

The N-terminal 200 amino acid residues of topoisomerase I (TopoN) is highly positive in charge and has DNA binding activity, without DNA sequence and topological specificity. Here, a fusion protein (6 x His-PTD-TopoN) containing a hexahistidine (6 x His) tag, a membrane penetration domain and TopoN (amino acid 3-200) was designed and developed. The protein can bind to different sizes (3.0-8.0 kb) and forms (circular and linear) of DNA and translocates the bound DNA to the nucleus. The protein also showed low cytotoxicity to GF-1 grouper fish fin cells that were previously very sensitive and difficult to transfect in vitro. Maintaining the hexahistidine tag increased the protein's transfection efficiency in COS7 African green monkey kidney cells and simplified the purification process. The plasmid pEGFP-N1 was delivered into COS7 cells by the protein in ATP- and temperature-dependent manners. The results indicate that the binding ability of TopoN is very useful for DNA delivery and the carrier protein can be expressed in Escherichia coli without removal of the hexahistidine tag.

• 出版日期2011-6