title: Induced binding of proteins by ammonium sulfate in affinity and ion-exchange column chromatography.
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.
authors.affiliation: Alliance Protein Laboratories, 3957 Corte Cancion, Thousand Oaks, CA 91360, USA. tarakawa2@aol.com
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Arakawa
authors.forename: Kouhei
authors.initials: K
authors.lastname: Tsumoto
authors.forename: Daisuke
authors.initials: D
authors.lastname: Ejima
authors.forename: Yoshiko
authors.initials: Y
authors.lastname: Kita
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Yonezawa
authors.forename: Masao
authors.initials: M
authors.lastname: Tokunaga
mesh_terms: D000645:Ammonium Sulfate; D000818:Animals; D002417:Cattle; D002846:Chromatography, Affinity / Q000379:methods*; D002852:Chromatography, Ion Exchange / Q000379:methods*; D011485:Protein Binding; D011506:Proteins / Q000302:isolation & purification*; D012685:Sepharose / Q000031:analogs & derivatives; D012710:Serum Albumin, Bovine / Q000302:isolation & purification
chemical_list: D011506:Proteins; D012710:Serum Albumin, Bovine; C029483:Blue-Sepharose; D012685:Sepharose; D000645:Ammonium Sulfate