1) it is possible that the tag was indeed lost (did you do an MS ?)
solution - use protease inhibitors all the way, and possibly change
expresison strains.
2) it is possible that the tag is not being exposed enough. Solution - move
to C-terminus, or introduce a Pro so that the tag is 'kinked'
A.G.E.
<mjcavallin at yahoo.com.ar> wrote in message
news:20030129173619.25968.qmail at ww02.hostica.com...
> I am working with the bacterial his tag expression system from Qiagen, and
am trying to purify the his-tag protein. Apparently, the recombinant protein
is going into inclusion bodies, so I am trying to purify it under denaturing
conditions (8 M urea). The problem is that after lysis there is a lack of
the His tag, because it is not longer recognised by Ni-NTA conjugate
(Qiagen) Does any one have any recommendations of how to generate a lysate
without losing the His tag?
>>>http://biowww.net/mynews/tree.php?group_name=bionet_molbio_proteins&begin=0
