| abstract |
A method is disclosed for the production of tissue factor pathway inhibitor (TFPI) which comprises expression of a non-glycosylated form of TFPI in E. coli as host and obtaining a highly active TFPI by in vitro refolding of the protein in which inclusion bodies isolated from the E. coli cells are subjected to stepwise purification comprising either series (A) or series (B) as follows:n (A)n (1) subjecting the inclusion bodies to sulfitolysis to form TFPI-S-sulfonate, (2) purifying TFPI-S-sulfonate by anion exchange chromatography, (3) refolding TFPI-S-sulfonate by disulfide interchange reaction, and (4) purifying active refolded TFPI by cation exchange chromatography, or (B)n (1) subjecting the inclusion bodies to reduction with ß-mercaptoethanol in urea to form reduced TFPI, (2) purifying the reduced TFPI by cation exchange chromatography, (3) refolding reduced TFPI by disulfide interchange reaction in urea, and (4) purifying the active refolded TFPI by cation exchange chromatography. |