I seem to recall a paper by the SGC group showing their flowscheme,
all proteins are first crystallized with the His-tagged variant, only
if no crystals are obtained or they diffract poorly a TEV cleaved
version is purified. I think the success rate of solved structures
between His-tagged and non-tagged was in the range of 70%. Sure one
could argue if they had cleaved off the tag to begin with and compared
both batches at the same time the ratio of His to non-His might be
different. Nevertheless it demonstrates that you don't need to cleave
of a His tag to be successful in obtaining a structure.
If I should find the paper again I will post the reference.
Jürgen
On 11 Nov 2008, at 06:34, [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
-
Jürgen Bosch
University of Washington
Dept. of Biochemistry, K-426
1705 NE Pacific Street
Seattle, WA 98195
Box 357742
Phone: +1-206-616-4510
FAX: +1-206-685-7002
Web: http://faculty.washington.edu/jbosch
