Objective：To construct a prokaryotic expression vector carrying thioredoxin peroxidase (TPx) gene of Branchiostoma belcheri Tsingtaunese and express it in E.coli.Methods: The cDNA fragments encoding TPx were obtained from Branchiostoma belcheri Tsingtaunese and were cloned into the expression vector pET-32a (+)M; the product was used to transform BL21(DE3) cells and expression of TPx protein was induced by IPTG.The recombinant TPx was expressed as a histidine fusion protein in E.coli and was purified with Ni chromatography and SP cation exchange chromatography.The expression and purification of TPx were analyzed by SDS-PAGE; the activity and structure of the protein were analyzed.Results: The recombinant plasmid pET-32a(+)M-TPx was highly expressed in E.coli.The purity of the protein was over 90% after purification; the molecular weight of the protein monomer was 24 460.The recombinant TPx protein existed as a mixture of both dimer and monomer.The recombinant TPx had a significant thiol-dependent peroxidase activity in the presence of dithiothreitol (DTT),and it could protect plasmid DNA and thiol-protein from damages caused by metal-catalyzed oxidation (MCO).Conclusion: The recombinant TPx protein has been successfully expressed in E.coli; its activity is closely related to the advanced structures.