Expression and purification of recombinant human hemangiopoietin in Escherichia coli⋆

ObjectiveTo express the soluble recombinant hemangiopoietin protein in E. coli BL21(DE3).MethodsUsing human fetal live cDNA as a template, a partial cDNA fragment of HAPO coding N-terminal region was subcloned into plasmids pTrc99, pQE60 and pET32c to construct different recombinant prokaryotic expression systems. After selecting, the soluble rhHAPO fusion protein was expressed stably in E. coli BL21 (DE3) by vector pET32c-HAPO and further isolated by nickelnitrilotriacetic acid (NTA) affinity chromatography. After cleavage with enterokinase, the rhHAPO protein was applied to Fast Flow SP sepharose column.ResultsThe rhHAPO protein had a purity of more than 95% and a good bioactivity based on the cell adhesion assay in ECV304 cells.ConclusionWe have established a protein engineering system to produce rhHAPO which may provide the possibility for clinical application.