Induced binding of proteins by ammonium sulfate in affinity and ion-exchange column chromatography |
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| Institution: | 1. Alliance Protein Laboratories, 3957 Corte Cancion, Thousand Oaks, CA 91360, United States;2. Department of Medical Genome Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Kashiwa, Chiba 277-8562, Japan;3. Amino Science Laboratories, Ajinomoto Co., Inc., Kawasaki, Kanagawa 210-8681, Japan;4. Department of Pharmacology, KEIO University School of Medicine, Tokyo 160-0004, Japan;5. Applied and Molecular Microbiology, Faculty of Agriculture, Kagoshima University, Kagoshima 890-0065, Japan;1. Department of Chemical Engineering, National Taiwan University of Science and Technology, 43, Keelung Road, Section 4, Taipei 10607, Taiwan;2. Department of Chemical Engineering, Cantho University, 3-2 Street, Cantho City, Viet Nam;1. College of Materials Science and Engineering, Sichuan University, No. 24, South 1st Section, 1st Ring Road, Chengdu 610065, China;2. The Military General Hospital of Chengdu, PLA, Tianhui Town, Chengdu 610083, China;1. Instituto de Biologia Experimental e Tecnológica, Apartado 12, 2781-901 Oeiras, Portugal;2. Instituto de Tecnologia Química e Biológica, Universidade NOVA de Lisboa, 2780-157 Oeiras, Portugal;3. LAQV-REQUIMTE, Departamento de Química, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, 2829-516 Caparica, Portugal;4. Departamento de Química, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, 2829-516 Caparica, Portugal;5. Sartorius Stedim Biotech GmbH, Spindler-Strasse11, 37079 Gottingen, Germany;1. National Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beijing 100190, PR China;2. University of Chinese Academy of Sciences, Beijing 100049, PR China;3. China Institute of Veterinary Drug Control, Beijing 100081, PR China;1. School of Chemical Engineering, Sungkyunkwan University, 2066 Seobu-ro, Jangan, Suwon 440-746, Republic of Korea;2. Department of Chemical and Biological Engineering, University of British Columbia, Vancouver, BC, Canada V6T 1Z3 |
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| Abstract: | In general, proteins bind to affinity or ion-exchange columns at low salt concentrations, and the bound proteins are eluted by raising the salt concentration, changing the solvent pH, or adding competing ligands. Blue-Sepharose is often used to remove bovine serum albumin (BSA) from samples, but when we applied BSA to Blue-Sepharose in 20 mM phosphate, pH 7.0, 50%–60% of the protein flowed through the column; however, complete binding of BSA was achieved by the addition of 2 M ammonium sulfate (AS) to the column equilibration buffer and the sample. The bound protein was eluted by decreasing the AS concentration or by adding 1 M NaCl or arginine. AS at high concentrations resulted in binding of BSA even to an ion-exchange column, Q-Sepharose, at pH 7.0. Thus, although moderate salt concentrations elute proteins from Blue-Sepharose or ion-exchange columns, proteins can be bound to these columns under extreme salting-out conditions. Similar enhanced binding of proteins by AS was observed with an ATP-affinity column. |
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