precipitation by ammuniom sulfate - (May/27/2011 )

I would like to precipitate entire protein content of supernatant for further examinations. Activity of proteins after re-suspending is important. I need some advise in the following topics with attention to the above mentioned concerns. Any help would be fully appreciated.

<*>Should I use the highest concentration of ammonium sulfate gradually in one step to precipitate all proteins or it may has adverse effect on low soluble bigger proteins activity?
<*>Regarding to save protein activity, which one is preferred? adding solid A. S. or saturated solution.
<*>Whether solid ammonium sulfate denature proteins or not?
<*>Is this method able to precipitate small peptides like cyclic dipeptides?
<*>Is it able to precipitate lipo-peptides and peptidoglycans too?

Regards - Hadi

Hadi on Fri May 27 08:14:34 2011 said:

<*>that's how i would do it. add a.s. slowly, with cooling and gentle stirring and allow to precipitate, in cold room or on ice for some time to enhance precipitation.<*>

<*>i've used both. using solid will avoid very large increases in volume. using a.s. solution to get to high concentrations will cause large increases in volume.<*>

<*>if added slowly and gradually then it won't be any worse than adding solution<*>

<*>in general, no<*>

<*>it should. we us it for glycoproteins without any problems.you may want to have a look at this protocol from cold springs harbor: