Hi Jorge,
using a system where you can cleave off the His-tag (with TEV, FactorX,
etc) adds a purification step which is complementary to the first
purification step (Ni-column etc). In my experience this results in very
pure protein which makes it more likely to crystallise.
Therefore I would always choose such a system and not spend much time on
trying to crystallise the protein with the His-tag attached.
Tim
--
Tim Gruene
Institut fuer anorganische Chemie
Tammannstr. 4
D-37077 Goettingen
GPG Key ID = A46BEE1A
On Tue, 11 Nov 2008, [EMAIL PROTECTED] wrote:
Dear all,
Concerning the crystallization of proteins with a His-tag, based
upon discussions on this bulletin board and on the article
Mike Carson, David H. Johnson, Heather McDonald, Christie Brouillette
and Lawrence J. DeLucas, His-tag impact on structure, Acta Cryst. D63,
295-301, (2007).
I understand that as a general approach one should try to
crystallize the protein with the His-tag (yet it might crystallize, so
no need to care the work of taking out the tag); then, if this is not
successful, go to the procedure to either express (and purify) it
without the His tag or take it out later. Any observations/advices?
But one other question to add is what if the protein is expressed
with both a c-myc tag and a His-tag (you might consider also the case
in which only the c-myc tag is present). Any experience on the effect of
the c-myc tag on crystallization? References are welcome yet I could not
find much googling around...
Thanks,
Jorge
