Expression and Purification of Zinc Finger Domain and Central Domain of MDMD2

[Objective] This study aimed to construct the His-tagged prokaryotic expression vectors harboring zinc finger domain(ZF) and central domain(acidic domain & zinc finger domain,CT) of MDM2,respectively,and preliminarily identified biologic activity of the purified fusion proteins.[Method]ZF and CT coding regions were amplified from MDM2 c DNA library by PCR and separately inserted into prokaryotic expression vector p ET28 b to construct the recombinant plasmids.After verification by enzyme digestion,the recombinant plasmids were separately transformed into E.coli DH5α competent cells.The expressed recombinant plasmids were purified using Ni-NTA magnetic beads and identified by SDS-PAGE and Western blotting.[Result]The molecular weight of His-ZF and His-CT fusion proteins was 21 and 31 k D,respectively.[Conclusion] Recombinant fusion proteins containing ZF and CT of MDM2 were obtained successfully,which laid the foundation for subsequent protein crystallization and three-dimensional structure analysis.