Expression and purification of truncated human recombinant nuclear apoptosis-inducing factor 1 in E.coli / 中国免疫学杂志

ABSTRACT

Objective:Nuclear apoptosis-inducing factor 1(NAIF1) is a novel apoptosis gene cloned in laboratory. To analyze the binding proteins of NAIF1 by pulldown method, the fusion expression vector of truncated human nuclear apoptosis-inducing factor 1〔NAIF1(73-327)〕 was constructed, were expressed and purified the recombinant GST-NAIF1(73-327) fusion protein in E.coli.Methods:The cDNA encoding human NAIF1(73-327) was amplified by PCR and cloned into pGEX-KG vector. The GST-NAIF1(73-327) fusion protein was expressed in E.coli and purified by affinity chromatography. The purified protein was detected by SDS-PAGE, Western blot and ESI-Q-TOF-MS/MS.Results:A prokaryotic expression vector of GST-NAIF1(73-327) was constructed and the GST-NAIF1(73-327) fusion protein was expressed in E.coli at high level. SDS-PAGE analyses indicated that the purified protein was about 53 kD. Western blot and MS/MS analyses verified the recombinant fusion protein.Conclusion:An efficient method for obtaining recombinant GST-NAIF1(73-327) fusion protein had been established and it could be used for further studies on the structure and function of NAIF1.