The effects of arginine on protein binding and elution in hydrophobic interaction and ion-exchange chromatography.
Arginine is effective in suppressing aggregation of proteins and may be beneficial to be included during purification processes. We have shown that arginine reduces non-specific protein binding in gel permeation chromatography and facilitates elution of antibodies from Protein-A columns. Here we have examined the effects of arginine on binding and elution of the proteins during hydrophobic interaction (HIC) and ion- exchange chromatographies (IEC) using recombinant monoclonal antibodies (mAbs) and human interleukin-6. In the case of HIC, the proteins were bound to a phenyl-Sepharose column in the presence of ammonium sulfate (AS) with or without arginine and eluted with a descending concentration of AS. While use of 1 M AS in the loading buffer resulted in complete binding of the mAb, inclusion of 1 M arginine in loading and equilibration buffer, only when using low-substituted phenyl-Sepharose, resulted in weaker binding of the proteins. While decreasing AS concentration to 0.75 M resulted in partial elution of the mAB, elution was facilitated with inclusion of 0.5-1 M arginine. In the case of IEC, arginine was included in the loading samples. Inclusion of arginine during binding to the IEC columns resulted in a greater recovery and less aggregation even when elution was done in the absence of arginine. These results indicate that arginine enhances elution of proteins bound to the resin, suggesting its effectiveness as a solvent for elution in HIC and IEC.
Arakawa
Tsutomu
T
Alliance Protein Laboratories, Thousand Oaks, CA 91360, USA.
Tsumoto
Kouhei
K
Nagase
Kazuo
K
Ejima
Daisuke
D
D000645:Ammonium Sulfate / Q000737:chemistry; D000911:Antibodies, Monoclonal / Q000737:chemistry; D001120:Arginine / Q000737:chemistry*; D002852:Chromatography, Ion Exchange; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D015850:Interleukin-6 / Q000737:chemistry / Q000302:isolation & purification; D011506:Proteins / Q000737:chemistry* / Q000302:isolation & purification*; D011994:Recombinant Proteins / Q000737:chemistry; D012997:Solvents
D000911:Antibodies, Monoclonal; D015850:Interleukin-6; D011506:Proteins; D011994:Recombinant Proteins; D012997:Solvents; D001120:Arginine; D000645:Ammonium Sulfate