Method for extraction and purification of hirudin mutant and use thereof

A method for extracting and purifying hirudin mutant HV2-Lys47 (SEQ ID NO.:

• 1) from a fermentation broth of Escherichia coli for fermentative production of hirudin mutant HV2-Lys47, comprising the steps of:
• treating the fermentation broth of Escherichia coli at an elevated temperature to sterilize the fermentation broth and remove impurity proteins;
• treating the sterilized fermentation broth by a ceramic membrane to remove the bacteria;
• treating the resultant fermentation broth from the ceramic membrane by an ultrafiltration membrane to remove impurity proteins;
• treating the resultant fermentation broth from the ultrafiltration membrane by a nanofiltration membrane to obtain a concentrated solution of crudely separated hirudin mutant HV2-Lys47;
• adding a salt into the concentrated solution of crudely separated hirudin mutant HV2-Lys47 as an auxiliary material, followed by spray drying to obtain dry powder;
• dissolving the dry powder in water and filtering out impurities;
• subjecting the filtrate to molecular sieve column chromatography only once, eluting the same with water, and concentrating the collected solution to obtain a crude hirudin mutant HV2-Lys47, wherein the molecular sieve column chromatography is selected from the group consisting of dextran gel G-25, dextran gel G-50, dextran gel G-75, and dextran gel G-100;
• and
• dissolving the crude hirudin mutant HV2-Lys47 in water, adding an organic solvent thereto to precipitate the hirudin mutant HV2-Lys47, and vacuum drying the same to obtain a pure hirudin mutant HV2-Lys47,
• wherein the organic solvent is selected from one or more of ethanol, methanol, acetone, or isopropanol, and the volume of the organic solvent added is 5-9 times volumes of the aqueous solution in which crude hirudin mutant HV2-Lys47 is dissolved.