Hsp70 is very hydrophobic, and the problem mentioned by you is quite
common in many affinity based purifications. I suggest that you load your
sample on an HIC (say, phenyl sepharose) and elute with a gradient from
high-salt to no-salt. Hsp70 typically elutes close to no-salt buffers on
the HIC chromatography.
Shekhar Mande
On Fri, 9 Mar 2007, Yong Tang wrote:
RE: Removal of bacterial chaperone Hsp70 contaminant from recombinant
protein preparation
Dear all,
I have a protein expressed at 37C for 3 hours in BL21 DE3 and purified
with sub-stoichiometric amount of apparent Hsp70 contaminant even
after exhaustive affinity (GST-fusion or His-tagged), ion-exchange and
sizing column steps. I would like to know if you have a
well-established protocol for getting rid of such a contaminant. I
asked around and was told to try adding ATP at certain(?) stage of the
purification.
I would much appreciate your input.
Many thanks! –yong @ the Wistar Institute
