Hi,
I am having a tough time trying to purify his-tagged proteins using
QUIAGEN system. First of all, the expression levels I am getting are
quite low (<200µg/L), but still, the protein is >90% in inclusion
bodies. When we purify this protein under denaturing conditions, we see
contaminant bands at an higher molecular weight. Finally, almost all
the recovered protein stick to dialysis tubing and cannot be recovered
at the final stage.
Does somebody have nice tricks to try in order to get a protein in the
soluble fraction ? Did somebody ever noticed the presence of other
bands on a Coomassie gel ? What can I do to increase the yield ? Any
idea of how I can do to avoid the protein to stick to the tubing ?
I will be most grateful if somebody can answer to one of more of my
problems. Thank you a lot !
Anne-Claude (gingras at medcor.mcgill.ca)
