Cloning,expression and purification of a new predicted virulence protein Z3672 of typeIII secretion system

Objective:To express and purify the new predicted virulence protein Z3672 of EHEC O157∶H7.Methods:The coding sequence of z3672 was amplified by PCR.The resultant fragment was sequenced and completely matched with the z3672 sequence of GenBank.It was cloned into the vector of pET-24a(+) to construct the recombinant plasmid pET-24a-z3672.The recombinant E.coli BL21(DE3)/pET-24a-z3672 was induced by IPTG,and the fusion protein was detected by Western blot and LC-MS/MS identity.Then the fusion protein was purified by washing the inclusion body.Results:Recombinant plasmid pET-24a-z3672 was expressed in E.coli BL21(DE3),and Z3672-His6 was purified by washing the inclusion body to a purity over 90%.Western blot and LC-MS/MS analyses showed that the fusion protein was expressed.Conclusion:The new predicted virulence protein Z3672 of EHEC O157∶H7 is successfully obtained and can be applied to further studies of the function of this protein and to further research on pathogenesis of EHEC O157∶H7.