Process for extracting and purifying the recombinant Placental Growth Factor and Placental Growth Factor in active form

摘要

Process for extracting and purifying the recombinant Placental Growth Factor (PLGF) in dimeric and multimeric active form containing no more than 1.5 % of monomeric form, by expression system in modified prokaryotic cells comprising the following steps: (I) fermentation of prokaryotic cells; (II) extraction and purification of the inclusion bodies; (III) renaturation of the expressed protein; (IV) ion-exchange chromatography; (V) reverse-phase chromatography, wherein step (I) is performed until obtaining optical density of the culture medium from 14 to 50 at 600 nm (OD600 14-50), followed by induction, step (II) comprises lysis of the culture cells, rupture of the DNA and isolation of the inclusion bodies, step (III) comprises solubilization of the inclusion bodies in denaturing buffer, whereby the expressed protein is brought at least partially back to the dimeric form, and in step (IV) the dimeric and multimeric forms of the expressed protein are separated from the monomeric form by an anion exchange chromatography, and in step (V) the dimeric and multimeric forms of the expressed protein are further separated from the monomeric form. Described is also Placental Growth Factor in active form obtainable by described process.