Prokaryotic expression and purification of chemokine CXCL16

LIN Meixiao; JIANG Yemei; CHEN Yaofeng; YANG Hua; XUN Yang

Abstract：To investigate the expression and purification conditions of transmembrane(TM-CXCL16)and soluble(sCXCL16)chemokine CXCL16,and to obtain CXCL16 protein with high expression yield and purity,this study amplified the gene fragments of TM-CXCL16,6×HisTEV-sCXCL16,and sCXCL16-6×His via PCR.These fragments were cloned into the pRSETA0 recombinant expression vector and transformed into the E.coli DH5α expression system,followed by induction with IPTG.Protein expression forms were identified by means of SDS-PAGE and Western blot,and proteins were purified by immobilized metal ion affinity chromatography(IMAC).Results showed that TM-CXCL16 and sCXCL16 proteins were successfully expressed in E.coli DH5α.Optimal expression conditions for TM-CXCL16 were 20 ℃,200 rpm,and 1.0 mM IPTG.Both sCXCL16-N(His-tagged at the N-terminus)and sCXCL16-C(His-tagged at the C-terminus)were expressed after IPTG induction,with the position of the His-tag showing no significant effect on sCXCL16 expression.Expression of sCXCL16-C was slightly higher at 20 ℃ compared to 37 ℃.Furthermore,IPTG concentration(0.1～1.0 mM)had no significant impact on sCXCL16-C expression levels.Analysis of expression forms revealed that sCXCL16-C was expressed in an insoluble form.The results indicate that both TM-CXCL16 and sCXCL16 were successfully expressed in a prokaryotic expression system.However,the sCXCL16 protein was expressed in an insoluble form,and high purity of the sCXCL16-C protein was not achieved using IMAC technology.Further optimization of the elution conditions is required to improve the protein purity.