Expression and Purification of Hi-lys Peptide,a Recombinant Relevant to Hirudin / 中国生物化学与分子生物学报

ABSTRACT

A new fusion expression vector, pED-P8-Hi-lys was designed and constructed. It includes four parts, a 20 peptide sequence of hirudin that can maintain anticoagulant activity, the C-terminus of asparaginase as a fusion partner, basic octopeptide (KRKRKKSR) that makes the fusion partner easy to remove, and the unique acid-labile aspartyl-prolyl bond. It was transformed into E. coli BL-21 and the fusion protein (AnsB-C-P8-Hi-lys) was expressed effectively as inclusion bodies after inducing by lactose. The objective peptide Hi-lys was purified by means of cell disruption, washing, ethanol precipitation, acid hydrolysis, and DEAE-cellulose 52 column chromatography. The antithrombin activity of the purified Hi-lys peptide was about 50 ATU/mg by thrombin activity assays.