The polybasic head indicative of your protein to have some possible membrane association? Try a detergent in your buffers might help. Also some of the polybasic head for protein need lipid association to be happy (PI and PCh)? Padayatti
On Fri, Aug 23, 2013 at 6:27 PM, Jahan Alikhajeh <ja...@graduate.org> wrote: > Dear Friends, > > I have been trying to purify a protein (27 kDa) which has a sumo plus 6 > His-tag at N-ter giving totaly a 45 kDa protein. > This protein does not express without sumo tag and has a poly basic tail > (Arg and Lys) at its N-ter. It does polymerize at acidic pHs. > When I tried to purify it with chelating Ni-NTA, it did not bind to the > column. I thought perhaps His-tag hided somewhere in the protein and is not > accessible thus, I repeated the experiments at 8 M urea. It did not make > any difference; very low binding to the column with high amount of unbound > protein in FT. Your advice is highly appreciated. > > Regards, > Jahan > > > -- PSP
