Surfactant precipitation and polar solvent recovery of α-chymotrypsin and ribonuclease-A

✍ Scribed by Youn-Ok Shin; Eva Rodil; Juan H. Vera

• Publisher: Elsevier Science
• Year: 2004
• Tongue: English
• Weight: 107 KB
• Volume: 17
• Category: Article
• ISSN: 1369-703X
• DOI: 10.1016/s1369-703x(03)00144-x

✦ Synopsis

The purification of two enzymes, ␣-chymotrypsin and ribonuclease-A, was studied using sodium di-(2-ethylhexyl) sulfosuccinate (AOT) as precipitating ligand. The enzymes formed a water insoluble complex upon contact with AOT and precipitated from the aqueous solution. The molar ratio between AOT and the targeted enzyme, required to obtain a 100% precipitation from an aqueous solution, was 7 for ␣-chymotrypsin and 13 for ribonuclease-A. Acetone was then used to recover the enzymes from the protein-AOT insoluble complex. The enzyme precipitated as white solid, while the surfactant remained in the acetone phase. The recovered enzyme was free of surfactant and retained its original enzymatic activity. The selectivity of this precipitation method was a strong function of the isoelectric point (pI) of proteins indicating that the ionic interactions between oppositely charged protein and AOT is the driving force for precipitation. Proteins with pI values within one pH unit, cannot be selectively separated using this method.