Posted: May 22nd, 2023

Blue-Sepharose and LDH as a model protein

A study was carried out by Arakawa et al to verify the improvement in the performance of column chromatography when arginine is utilized in the dye affinity chromatography (410). The dye-affinity column chromatography provides a faster and more convenient protein purification protocol. This is because proteins usually have a high affinity for dye columns (Arakawa et al. 2007). In addition, a bigger proportion of samples can be eluted without preceding treatment procedures. In this method, “covalently attached group-specific ligands are used for purification of enzymes, such as dehydrogenases and kinases” (Arakawa et al. 2007).

Columns with blue dye adhere to enzymes that need “adenyl-containing cofactors such as NAD and NADP. Lactate dehydrogenase (LDH) binds to blue dye columns and the bound enzyme is eluded by salts, but with low recovery and broad elution peak ” (Arakawa et al. 2007). In this study, arginine was investigated to find out whether it can solve these challenges. “Thus its ability to increase collection and resolution using Blue-Sepharose and LDH as a model protein” (Melander. et al. 1999)

The results of the study revealed that the usual eluent, “NaCl indicated a wide elution peak with a low recovery of lactate dehydrogenase, at most approximately 60 using 2M salt The recovery was seen to decrease with either increase or decrease in the NaCl concentration” (Arakawa et al. 2007) However, when arginine was used in place of NaCl, the recovery of lactate dehydrogenase was higher than 80% at more than 0.5 M (Arakawa et al. 2007). The recovery was almost free of the arginine concentration used (Mattiasson et al. 1999). This resulted in a “sharper elution peak, leading to the recovery of a protein solution of good concentration” (Arakawa et al. 2007).