Arginine improves protein elution in hydrophobic interaction chromatography. The cases of human interleukin-6 and activin-A.
The effects of arginine on protein binding and elution in hydrophobic interaction chromatography (HIC) were examined using recombinant human interleukin-6 (IL-6) and activin-A. Binding of IL-6 in the presence of ammonium sulfate (AS) was tested using low- and high-substituted phenyl-sepharose. While inclusion of arginine during loading of IL-6 resulted in incomplete binding to the low-substituted phenyl-sepharose, binding was complete to the high-substituted phenyl-sepharose. Arginine facilitated elution of IL-6 from both columns. These results demonstrate that arginine weakens hydrophobic interactions between IL-6 and the phenyl-sepharose. More drastic results were obtained using activin-A, which showed undetectable recovery from phenyl-sepharose. Although no apparent elution of activin-A was observed from butyl-sepharose in aqueous buffer alone, the addition of arginine to the buffer resulted in partial elution recovery and, together with ethanol, resulted in greatly improved recovery of the protein. Two arginine derivatives, acetylarginine and agmatine, were also effective. These results show that arginine improves protein elution in HIC.
Tsumoto
Kouhei
K
Department of Medical Genome Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Kashiwanoha, Kashiwa 277-8562, Japan. tsumoto@k.u-tokyo.ac.jp
Ejima
Daisuke
D
Nagase
Kazuo
K
Arakawa
Tsutomu
T
D028341:Activins / Q000302:isolation & purification*; D001120:Arginine / Q000737:chemistry*; D002853:Chromatography, Liquid / Q000379:methods*; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D015850:Interleukin-6 / Q000302:isolation & purification*; D011485:Protein Binding / Q000187:drug effects; D011994:Recombinant Proteins / Q000302:isolation & purification; D012685:Sepharose / Q000031:analogs & derivatives
D015850:Interleukin-6; D011994:Recombinant Proteins; C422405:activin A; D028341:Activins; C020927:phenyl-sepharose; D012685:Sepharose; D001120:Arginine