Prokaryotic Expression and Biological Activity of Poly-Arg Protein Transduction Domain-Apoptin Fusion Protein

Objective To express poly-Arg protein transduction domain(PTD)-apoptin fusion protein in prokaryotic cells and determine its biological activity. Methods Arg9-VP3 sequence was amplified by PCR and inserted into vector pET-43.1a, and the constructed recombinant plasmid pET43.1a-Arg9-VP3 was transformed to E. coli BL21(DE3)for expression under induction of IPTG. The expressed product was purified by Ni2+-NTA chromatography, lysed with enterokinase, concentrated by ultrafiltration and determined for biological activity. Results Both restriction analysis and sequencing proved that recombinant plasmid pET43.1a-Arg9-VP3 was constructed correctly. Recombinant fusion protein was expressed in a soluble form, reached a purity of more than 90% after purification and showed inhibitory effect on the proliferation of HeLa cells. Conclusion Poly-Arg PTD-apoptin fusion protein was successfully expressed in E. coli and purified , which induced the apoptosis of HeLa cells.