PROCESS FOR MANUFACTURE OF A MODIFIED COLLAGEN-INDUCED PLATELET AGGREGATION INHIBITOR PALLIDIPIN

摘要

The invention provides a process for manufacture of a recombinant protein called Asp-Pallidipin. Asp-Pallidipin inhibits the collagen-induced platelet aggregation of mammalian platelets. The Asp-Pallidipin comprises: (i) a protein (Pallidipin) selected from the group of Pallipin proteins, and (ii) the amino acid aspartic acid, wherein the aspartic acid is connected by a peptide bond with the N-terminal end of Pallidipin. The process comprises the steps aa) transfecting at least one bacterium with an appropriate vector, wherein the vector comprises: (i) a DNA or cDNA coding for the recombinant Asp-Pallidipin, (ii) a suitable signal peptide sequence which signal sequence is cleaved so that the amino acid aspartic acid is in the position +1 of the amino acid sequence seen from the position of the Pallidipin and (iii) a suitable promoter; bb) expressing the preprotein comprising Asp-Pallidipin and the signal sequence; cc) transporting the Asp-Pallidipin from the cytoplasm of the bacterium to the periplasm, cleavage of the preprotein by at least one protease during the transport, producing the Asp-Pallidipin, dd) isolating the Asp-Pallidipin by extracting the periplasm, and ee) purifying the Asp-Pallidipin. The protein is used as a medicament for inhibiting of collagen-induced human platelet aggregation or of cancer with metastatic tumor cells.