I had a similar case before. I could not find notes for the details. 1. During the lysis, add protein amine sulfate and spin down with the speed up to 10K rpm (normally 8k) for 30 mins. 2. adding polysaccharide (1~ 5 %) as additive to adjust nonspecific bind. For polysaccharides, I tried several kinds of them ordered from Sigma. I forgot which one was the best (sorry). 3. use ionic columns ( anionic + cationic columns, chained them together) at 4C. 4. Then, used Co-Column (not, NTA,) for further purification. ...
The final purity could reach 95%. However, l lost a lot of protein during purification. On Fri, Sep 15, 2017 at 4:53 AM, Narayanan Ramasubbu < [email protected]> wrote: > Hi. We are working on a periplasmic protein that breaks naked glycans in > peptidoglycans. There is truncated structure available but our target is > the full length protein. The difficulty us that it strongly binds to the > resin with or without his.tag. Changing the resin to acrylamide did not > help. > Has anyone come across similar problem and how was it resolved. > The pdb structure is the catalytic domain and mussing a region that, in my > opinion, binds to the resin. > Thank you in advance > Sent from my iPhone -- Kevin Jin Sharing knowledge each other is always very joyful...... Website: http://www.jinkai.org/
