[Expression and purification of recombinant hirudin].

OBJECTIVE

To express the recombinant hirudin variant 1 (rHV1) with biological activity in prokaryotic cells and then isolate and purify the expressed products.

METHODS

The hirudin variant 1 gene in plasmid vector pBV220 was expressed in E. coli strain DH5 alpha. The biological activity of rHV1 was determined with chromogenic substrate method. The expressed product was purified by ultrafiltration, DEAE-Sephadex A-50 filtration and thrombin-Sepharose 4B affinity chromatography.

RESULTS

The expressed rHV1 accounted for approximately 16.9% of E. coli cell proteins with an activity of 20-30 ATU (antithrombin unit) per milliliter culture. The purified rHV1 showed a homogeneous band on SDS-PAGE.

CONCLUSIONS

This is the first report in China on successful expression of hirudin variant 1 gene in E. coli and purification of the expressed rHV1.