Hello,
I would like to express a protein using a cell free protein expression kit from New England Biolabs. The protein is a chymotrypsin and so it must be activated through cleavage to be functional. I am trying to understand the best way to express this protein. I am not sure if I express only the sequence of the activated protein if that would cause problems (e.g. from my understanding the zymogen form can sometimes help fold properly). Alternatively, I could express the inactive form and hope it becomes activated in my assay (I will be applying it to some tissue in which it is normally found). Or, since chymotrypsins are typically activated by cleavage by trypsin, I could perhaps incubate with tagged trypsin first (and remove trypsin using affinity purification). I have no experience with expressing proteins so any thoughts on what would work best are greatly appreciated. Thanks!
Jeremy
