[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.
Institute of Hematology, CAMS and PUMC, Tianjin 300020.
J
J
Guo
H
H
Wang
J
J
Gu
D004926:Escherichia coli / Q000235:genetics; D005343:Fibrinolytic Agents / Q000302:isolation & purification; D015870:Gene Expression; D014644:Genetic Variation; D006629:Hirudins / Q000096:biosynthesis* / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification
D005343:Fibrinolytic Agents; D006629:Hirudins; D011994:Recombinant Proteins