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US 2010/0144622 A1Published application
Methods for production of recombinant plasminogen and plasmin polypeptides
OwnerXinli Lin, Daniel Medynskiinventors
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Abstract
Methods of producing properly refolded recombinant plasminogen and plasmin polypeptide are provided. Denatured recombinant plasminogen polypeptide is refolded by first solubilizing the polypeptide with a chaotroph and reducing and oxidizing agents at high pH, followed by refolding in the presence of reduced concentration of chaotroph and reducing and oxidizing agents and in the presence of arginine.
Claims25
A method for refolding a recombinant plasminogen polypeptide, comprising:
- (a) solubilizing a plasminogen polypeptide in a solubilization buffer, said solubilization buffer comprising a high concentration of chaotroph, a reducing agent, redox reagents, and having a pH of about 9.0 to about 11.0, thereby producing a solubilized plasminogen polypeptide solution;
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- (b) rapidly diluting said solubilized plasminogen polypeptide solution with a refolding buffer by adding said solubilized plasminogen polypeptide solution into the refolding buffer, thereby producing diluted solubilized plasminogen polypeptide solution, wherein the refolding buffer comprises arginine and has a pH of about 8.0 to about 10.0;
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- (c) incubating the diluted solubilized plasminogen polypeptide solution, thereby producing a refolded plasminogen polypeptide.
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