PROCESS FOR ACTIVATING HETEROLOGOUS, EUCARYOTIC PROTEINS GENETICALLY ENGINEERED AND PRESENTING DISULPHIDE BRIDGES AFTER THEIR EXPRESSION IN PROCARYOTIC CELLS

摘要

To activate heterologous eukaryotic proteins produced by genetic engineering and having disulfide bonds after expression in prokaryotes, the disintegration of cells, solubilization under denaturing and reducing conditions and their activation under oxidizing conditions in the presence of GSH / GSSG, or is used during the activation step a pH between 9 and 12, a GSH concentration of 0.1 to 20 mmol / l, a GSSG concentration of 0.01 to 3 mmol / l and a concentration not -dénaturante of the denaturing agent, or reducing agents are separated and denaturant, transforms the thiol groups of proteins in the mixed disulphides of protein and glutathione by addition of GSSG under denaturing conditions and used for the activation step a pH between 7 and 10.5, a GSH concentration of between 0.5 and 5 mmol / l and a non-denaturing concentration of the agent dénatur ant. This method is particularly useful for t-PA and beta interferon.