PRECIPITATION

Ammonium Sulfate Calculator: 

Ammonium Sulfate Precipitation is a simple and effective means of fractionating proteins. 

Information (refer this article for more information doi: 10.1002/0471140864.psa03fs13 )

The solubility of globular proteins increases upon the addition of salt (<0.15 M), an effect termed salting-in. At higher salt concentrations, protein solubility usually decreases, leading to precipitation; this effect is termed salting-out . Salts that reduce the solubility of proteins also tend to enhance the stability of the native conformation. In contrast, salting-in ions are usually denaturants.

The mechanism of salting-out is based on preferential solvation due to exclusion of the cosolvent (salt) from the layer of water closely associated with the surface of the protein (hydration layer). The hydration layer, typically 0.3 to 0.4 g water per gram protein , plays a critical role in maintaining solubility and the correctly folded native conformation. There are three main protein-water interactions: ion hydration between charged side chains (e.g., Asp, Glu, Lys), hydrogen bonding between polar groups and water (e.g., Ser, Thr, Tyr, and the main chain of all residues), and hydrophobic hydration (Val, Ile, Leu, Phe). In hydrophobic hydration, the configurational freedom of water molecules is reduced in the proximity of apolar residues. This ordering of water molecules results in a loss of entropy and is thus energetically unfavorable. When salt is added to the solution, the surface tension of the water increases, resulting in increased hydrophobic interaction between protein and water. 

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