I expressed a fusion protein using pet32a and purifing it by IMAC. But when this protein was cleaved by EK enzyme at temperature 23℃ for 16hr,the objected protein(about 15KD)was degradation to some little molecular weight .So i reduced the enzyme cleavage time to 2hr, the protein degradation still exists. I also try to change the cleavage temperature to 4℃,but it does not work. Could anyone please give me some idea and suggestions how to solve the protein deagradation after EK cleavage?
protein degradation after EK cleavage