Soluble expression and bioactivity analysis of THANK
Objective To construct THANK expressing vector and induce its protein expression in E.coli and to analyze its bioactivity. Methods The extracellular fragment of THANK was amplified by PCR using pMD18 THANK plasmid containing a full length THANK cDNA and the extracellular fragment was subcloned into the pET 11a vector resulting in a recombinant plasmid pET THANK, the plasmid was inducted with IPTG in different conditions and the expressed protein was analyzed and identified by SDS PAGE/Western blot. The function of purified THANK protein was identified. Results THANK extracellular cDNA was amplified by PCR, SDS PAGE. Western blot analysis showed that THANK protein was expressed. Further bioactivity assay indicated that the soluble protein can costimulate B, T cell proliferation and induced activated T cells apoptosis. Conclusion THANK extracellular cDNA was successfully cloned and expressed. The bioactivity analysis showed that the protein was functioning in the modulation of immune response.