Human leukotriene C4 synthase (PDB accession code: 2UUI) is another
example, illustrating how an N-terminal polyhistidine-tag, in
conjunction with metals, presumably facilitated crystallization.
On Jun 27, 2012, at 12:04 PM, Brad Bennett wrote:
I think it was an N-terminal RGS-type His tag in 3O8Y (human
lipoxygenase) that mediated crystal contacts with a symmetry related
molecule. As I recall, this tag composed a B-strand that formed a
nice interface with a "native" B-strand of the symmetry related
molecule. Pretty cool...
-Brad
On Wed, Jun 27, 2012 at 11:00 AM, Phoebe Rice <pr...@uchicago.edu>
wrote:
With Flp recombinase - DNA complexes, a C-terminal His tag triggered
a different (but sadly not better) crystal form, and the His side
chains packed against the bases at the end of a neighboring DNA
duplex.
=====================================
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
phone 773 834 1723
http://bmb.bsd.uchicago.edu/Faculty_and_Research/01_Faculty/01_Faculty_Alphabetically.php?faculty_id=123
http://www.rsc.org/shop/books/2008/9780854042722.asp
---- Original message ----
>Date: Wed, 27 Jun 2012 10:14:58 -0400
>From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> (on behalf of "R.
M. Garavito" <rmgarav...@gmail.com>)
>Subject: Re: [ccp4bb] The effect of His-tag location on
crystallization
>To: CCP4BB@JISCMAIL.AC.UK
>
> Most of the comments you will get will be anecdotal
> in that people will report the successful results
> and do not take the time or effort to characterize
> the less successful results. This often occurs
> because the tagged portion of the protein is most
> often disordered, even in the best crystals. Thus,
> other than saying "tagging on this end works, but
> tagging on that end doesn't," there is little more
> you can say. Each case will be different, and it is
> almost impossible to arrive at any generalized
> conclusion.
> We prefer C-terminal tagged proteins for a number of
> reasons, but if an N-terminally tagged protein
> crystallizes well, so be it. Of the dozens of N-
> and C-tagged protein structures we have solved in my
> lab and with collaborators, I have only seen one
> case of an ordered His-tag: the His residues had
> coordinated Cd ions, which proved essential for
> getting good crystals. However, beyond that there
> was not much more to say.
> For your protein and the resulting crystals, an
> N-terminally tagged protein crystallized well.
> Whether you can draw any more conclusions from
> these results depends on characterizing crystals of
> both N- and C-tagged proteins. Just assuming that
> the C-tagged protein is trying to crystallize in the
> same or related crystal form as the N-tagged protein
> is an unwarranted assumption without experimental
> evidence to back it up. That is why most groups
> just run with the winner.
> Cheers,
> Michael
> ****************************************************************
> R. Michael Garavito, Ph.D.
> Professor of Biochemistry & Molecular Biology
> 603 Wilson Rd., Rm. 513
> Michigan State University
> East Lansing, MI 48824-1319
> Office: (517) 355-9724 Lab: (517) 353-9125
> FAX: (517) 353-9334
> Email: rmgarav...@gmail.com
> ****************************************************************
> On Jun 26, 2012, at 9:06 PM, weliu wrote:
>
> Dear all,
>
> We crystallized a protein and found that crystal
> quality greatly depended on the location of
> His-tag. When a His-tag was added at the
> C-terminus, only crystalline precipitate or
> spherical quasi crystals were grown. However, when
> the His-tag was moved to the N-terminus, single
> crystals were grown under a number of conditions,
> and the best one diffracted to 1.7 angstrom after
> optimization. I was wondering if there were
> published reports describing similar cases.
>
> Thank you in advance
>
> Wei Liu
