High-level synthesis of biologically active human plasminogen activator inhibitor type 1 (PAI-1) in Escherichia coli |
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Authors: | W P Sisk G L Davis D Kingsley A T Chiu T M Reilly |
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Affiliation: | E.I. DuPont de Nemours and Co., Central Research and Development Department, Wilmington, DE 19880. |
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Abstract: | Segments of a cDNA encoding human plasminogen activator inhibitor type 1 (PAI-1) were subcloned into a highly regulated and inducible Escherichia coli expression system. A plasmid encoding the mature form of human endothelial PAI-1 produced a functional recombinant molecule, as indicated by its ability to inhibit tissue plasminogen activator's enzymatic activity. In contrast to PAI-1 isolated from human fibrosarcoma cells, the biological activity of the recombinant PAI-1 was not dependent on pretreatment with denaturing agents. A construct encoding a polypeptide lacking the first 80 amino acids of PAI-1 also produced elevated levels of the truncated recombinant protein. However, this truncated product was functionally inactive, indicating that an intact N terminus is required for activity. |
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