Soluble Expression,Purification and Characterization of Ad5-knob Protein

Ad5-knob gene from AdEasy plasmind DNA was amplified by polymerase chain reaction (PCR) and cloned into an E. coli expression vector pQE30 digested with BamH I and Hind III. After induction with IPTG, the transformed E. coli strain M15 expressed Ad5-knob gene efficiently. Moreover, the recombinant Ad5-knob protein mainly existed as a soluble protein. After one step purification with Ni~(2+)-NTA affinity chromatography, the protein was purified to nearly homogeneity. N-terminal amino acid sequencing indicated N-terminus of purified Ad5-knob. When FITC-labeled Ad5-knob transfers Hela cells, it can specially bind to the receptor to the surface of Hela cells and enter the cytosol. It suggests that the recombinant Ad5-knob protein identified and purified is to be suitable for nonviral gene carriers as a target protein in future studies.