Storing proteins - (Jun/08/2006 )
What is the significance of keeping proteins at 4 degrees in experiments? Why is thawing and freezing again at 20 degrees so bad for the proteins?
protein are kep at 4 degrees during experiments to limit the effect of proteases, degradation etc as most of these do not occur at lower tempratures.
freeze thaw cycles affect protein activity, this is due to the water shell surrounding proteins interacting differently with the protein than under normal circumstances (above 0 degrees), once or twice and you wont be able to tell the difference but numerous freeze thaw cycles and the protein begind to lose its biolocial activity.
In the end most natural proteins have not evolved to be resistant/unaffected against freeze thaw cycles as they do not occur naturally, thus they are volatile under these conditions
Actually, I find that many proteins (without known enzymatic activity) are greatly affected by freeze thaw cycles. In particular, proteins that precipitate readily are more likely to precipitate if left at warmer temperatures (4 degrees, -20 degrees) or if there are multiple freeze-thaw cycles. To avoid this, we aliquot all of proteins to multiple tubes and generally store at -80degrees. Proteolysis is also affected. I do not not know the mechanism for increased precipitation, but I assume proteins are more mobile in solution which may allow protein-protein association and eventually insolubility. I am not sure, though.