His6-tagged protein purification

Angela Manning a.manning at ualberta.ca
Wed Feb 23 14:03:17 EST 2000


I am trying to purify a bacterially exressed His6 tagged protein on a
Probond column under native conditions.  I have purified the protein under
denaturing conditions, however, the protein is "completely" insoluble after
denaturation.  So, I want to purify the soluble fraction under native
conditions.  However, when I do this the protein is contaminated with lots
of other proteins, even after multiple washes.  I have also tried washing
in a low concentration (40mM) of imidazole but my protein largely washed
off under these conditions. Does anyone have any suggestions as to how I
might purify this protein?  Thank you!

-- 
Angela Manning       a.manning at ualberta.ca
Dept of Biological Sciences
University of Alberta
Edmonton, AB Canada
T6G 2H3




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