Protease or Nuclease Contamination

A more gentle method of dehydration is the dropwise addition of a saturated ammonium sulfate solution whose pH has been previously adjusted. This is usually the method of choice for precipitation of purified or partially purified proteins in small volumes since the rate of protein dehydration can be carefully controlled. Unfortunately, this method is undesirable for large volumes, because ammonium sulfate addition 

In practice precipitates are usually collected by centrifugation at intervals such as those shown in Table 10-1. In this case most of the enzyme activity is found in the 35 to 45% and the 45 to 55% precipitates. These data suggest that an initial fractionation of 0 to 35% and then a 

Percentage (NH lzSO (saturated at 25°C, 4.1M) Activity Precipitated (arbitrary units) 

There are two major advantages of ammonium sulfate fractionation. First it is usually possible to discard up to 75% of the crude protein at this 

The organic solvents most often used to bring about precipitation include ethanol, methanol, and acetone. Dioxane and tetrahydrofuran are adequately soluble in water, but are usually contaminated with peroxides which are quite deleterious to most proteins. Hence these solvents are rarely used. 

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