Improved performance of column chromatography by arginine: dye-affinity chromatography.
Arginine has been effectively used in various column chromatographies for improving recovery and resolution, and suppressing aggregation. Here, we have tested the effectiveness of arginine as an eluent in dye-affinity column chromatography using Blue-Sepharose, which binds enzymes requiring adenyl-containing cofactors (e.g., NAD). A common eluent, NaCl, showed a broad elution peak with low recovery of lactate dehydrogenase, at most approximately 60% using 2M salt. The recovery decreased as the NaCl concentration was either decreased or increased; i.e., the recovery was maximum at 2M. On the contrary, addition of arginine to the eluent resulted in more than 80% recovery above 0.5M and the recovery was nearly independent of the arginine concentration. The elution peak was much sharper with arginine, leading to elution of more concentrated protein solution. Successful elution of proteins bound to the ATP-agarose resins by arginine was also described.
Arakawa
Tsutomu
T
Alliance Protein Laboratories, Thousand Oaks, CA 91360, USA. tarakawa2@aol.com
Ejima
Daisuke
D
Tsumoto
Kouhei
K
Ishibashi
Matsujiro
M
Tokunaga
Masao
M
D001120:Arginine / Q000737:chemistry*; D002846:Chromatography, Affinity / Q000379:methods*; D004396:Coloring Agents / Q000737:chemistry; D007770:L-Lactate Dehydrogenase / Q000737:chemistry / Q000302:isolation & purification; D012685:Sepharose / Q000031:analogs & derivatives* / Q000737:chemistry
C030548:ATP-sepharose; D004396:Coloring Agents; C029483:Blue-Sepharose; D012685:Sepharose; D001120:Arginine; D007770:L-Lactate Dehydrogenase