PROTEIN C PURIFICATION USING AQUEOUS TWO-PHASE SYSTEMS

THE OBJECTIVE OF THIS PROGRAM WILL BE TO USE AQUEOUS POLYMER TWO-PHASE SYSTEMS FOR THE PURIFICATION OF PROTEIN C FROM HUMAN PLASMA. BOTH PHASES CONTAIN GREATER THAN 80 PERCENT WATER WITH THE UPPER PHASE RICH IN POLYETHYLENE GLYCOL (PEG) AND THE LOWER PHASE RICH IN A POLYMER OR SALT. THE EXPERIMENTAL PROCEDURE WILL BE AIMED AT EXTRACTING PROTEIN C INTO THE TOP PHASE OF AN AQUEOUS TWO-PHASE SYSTEM THROUGH INTRODUCTION OF CHARGED, HYDROPHOBIC OR AFFINITY PEG DERIVATIVES WHICH HAVE A SPECIFIC ATTRACTION FOR PROTEIN C. THE REMAINING PLASMA PROTEINS WILL PARTITION TO THE BOTTOM PHASE. THE POTENTIAL MEDICAL APPLICATIONS FOR PROTEIN C INCLUDE NOT ONLY PEOPLE WHO ARE DEFICIENT IN THE PROTEIN C, BUT PEOPLE WHO MAY EXPECT TO HAVE BLOOD CLOTTING PROBLEMS BECAUSE OF ANTICIPATED SURGERY OR ORGAN IMPLANTATION, AND HEART ATTACK PATIENTS WITH BLOOD CLOTTING PROBLEMS. AQUEOUS POLYMER TWO-PHASE SYSTEMS PROVIDE SUBSTANTIAL BENEFITS OVER COMPETITIVE TECHNOLOGY SUCH AS LIQUID CHROMATOGRAPHY, HPLC AND ELECTROPHORESIS. THE POLYMERS AND BUFFERING SALTS WHICH COMPRISE THE SYSTEMS ARE NON-TOXIC AND HAVE BEEN SHOWN TO STABILIZE THE BIOLOGICAL ACTIVITY OF BIOMOLECULES. THE PHASE SEPARATION OCCURS UNDER GENTLE CONDITIONS AND CAN EASILY BE SCALED UP FOR LARGE-SCALE PURIFICATION.