I hope somebody can help me! I am isolating a protein with His-Tag.
In order to get rid of the poly-histidine I am doing a rTEV-protease
digestion. The problem is, that in the end of the isolation the protein
is in 4M Urea, 500 mM Imidazol, but the protein doesn't work with such
high Urea concentrations.
So I am trying to change the buffer by centrifugation with centricon
concentrators from amicon. But yield is only about 10%. The protein seems
to dissapear during the concentration procedure. I can find it neither
in the flow-through, nor in the retentate.
Thanks in advance
Please send e-mail to: werner at inw.agrl.ethz.ch
