Hi Narayanan,We had similar problems with a membrane protein. It did bind Ni-NTA resin strongly. We washed out the resin (Ni-NTA) with high imidazole to sealer all other proteins and eluted out protein with very small concentration of SDS at room temp with shaking. Please read our paper in- Huang et al., Biochemistry, 49, 1115-1126, 2010Hope this procedure can help you too.Smita
On Friday, September 15, 2017 6:54 AM, Narayanan Ramasubbu <[log in to unmask]> 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
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