Dear ALL;
One of our proteins will gain activities after cleavage between secondary
structures(probably at three sites). In the intact structure, the N-terminal
and C-terminal domains beside this cleavage site have extensive contacts. So
far we are stilling looking for the enzymes that cut this protein in human,
probably at disease situation.
However, when we made the truncates, both the N-ter and C-ter domains
become soluble aggregates even during expression at 20 degree, whereas the
whole enzyme can be solubly expressed in E.coli.
Now we are trying to design an artificial cleavage site, say TEV or thrombin
site, to at least prove the idea at the biochemistry level.
There are some short loops between strands and helices.
Can anyone suggest a good way of adding in the cleavage site without changing
the native structures?
Thanks a lot.
Jerry McCully
