protein denaturation

Question

write a simple technique that can be conducted in the laboratory to identify the effect of shaking and freezing on protein denaturation.

Answer 1

Shaking and freezing can alter the nature of the proteins. For instance, vigorous shaking can denature proteins and will result in precipitation. If this happens for any given protein then the optical density at 280 nm can be measured to see this difference. here is a reference- Reese and Robbins, Journal of Colloid and Interface Science, Vol. 83, No. 2, October 1981

For freezing, it is often said that the uncontrolled presence and formation of ice is what causes the proteins to crystallize thereby altering their 3D structure leading to denaturation. However freezing and thawing leads to denaturation followed by rapid degradation.

SDS-PAGE electrophoresis can also be conducted and the banding patterns of the proteins can be observed by coomassie blue dye staining. Non denatured proteins tend to have a smear pattern and the bands will be at very high molecular weight range (instead of a clear and sharp banding pattern) compared to denatured control and otherwise.

Additionally given these shaking and freezing conditions, performing an QC by a simple spectrometry analysis (Bradford method or BCA method) on those protein samples will provide an idea on the protein concentration and the overall nature. Degraded proteins will have a very low absorption spectra in this method compared to the control and sample which did not go through repeated thawing and freezing.