It does not look like protein crystal and IZIT staining is not reliable in
determining protein or other. Mostly it is like detergent or PEG crystal or
quasi-crystal.
Good luck
Wu
2010/8/31 qiangm zhang
> Hi all,
>
> I got a crystal of one membrane protein (~60kD) from Na/K phosphate
> co
When diffraction fails - one can always look at the crystals with a UV
microscope (assuming you have Trp) - they should light up. Or if its
possible - harvest them, wash and run a gel. (or if its detergent a TLC)
Joe
2010/9/1 wu donghui
> It does not look like protein crystal and IZIT stainin
On Wed, Sep 1, 2010 at 4:26 AM, Ed Pozharski wrote:
> The
> reason you see the missing region in (2mFo-DFc) map is because it is
> effectively the sum of model map (mFo) which shows you the parts of the
> model you have already placed and difference map (mFo-DFc) which shows
> you the regions whic
Dear CCP4,
In our automated data processing and refinement pipeline, phaser
sometimes comes up with solutions in non-standard settings (e.g. P 21 2
21). These solutions subsequently fail in autobuster and it turned out
that this is because autobuster invokes sftools and sftools apparently
is not a
Dear Herman,
please find attached a patch for sftools that will add support
for the following non-standard space group settings:
A2, C21, I21, P2122, P2212, P21221, P22121
Note: the number for I21 follows the upcomming change to
syminfo.lib: 3004 -> 5005.
diffstat -p0 <
CCP4-20091104-src-sft
I cannot say what you have - running a gel, MS, etc of a washed crystal
could confirm what you have.
What you did not indicate is if your lack of diffraction was of frozen
or unfrozen crystals. I have seen too many cases where it is the cryo
condition killing diffraction. So if you have not tried
I cannot say what you have - running a gel, MS, etc of a washed crystal
could confirm what you have.
What you did not indicate is if your lack of diffraction was of frozen
or unfrozen crystals. I have seen too many cases where it is the cryo
condition killing diffraction. So if you have not tried
Dear Claus,
Thank you very much for this patch. We will install it, and I hope CCP4
will install it quickly as well ;-). Still I do not understand why
sftools has all symmetry operations hardcoded, while most other programs
use the CCP4 libraries. In that way, sftools would always be up to date
an
Ian is, as always, absolutely right. The only comment/correction I have
is that Hailang was apparently referring to severely incomplete model,
for which the poor phases will dominate the mFo map. Under such
circumstances, even 2fo-fc map will not correctly reflect the actual
relative contribution
Following Ian's excellent comment, do I understand correctly that
2mFo-DFc is the maximum likelihood estimate of the full model map (i.e.
the best map possible) or it's simply modification of 2Fo-Fc map where
plain Fo/Fc are replaced by their maximum likelihood estimates? Other
words, is k=2 the m
Hi Herman,
As a former biomol user you might have guessed why. SFTOOLS found its
origin as a transitional program helping the Groningen group move to the
CCP4 mtz format. Since the Groningen MDF and CCP4 mtz had different
ideas about space group symmetry and, especially, asymmetric unit
defin
I am trying to find a MR solution for a large unit
cell (R3:H, 158x158x196) with a relatively poor, but I think workable
search model (30% identity, 50% similarity). The data set is decent to
2.4 A. I might be able to get better if necessary. I submitted the
sequence of the target to the Phyre
Hi Roger,
personally, I would take the four dimers from Phaser, strip off all side
chains (make a poly-ala/gly), do a ML refinement with tight NCS
restraints. The resulting map could then be on-the-fly
real-space-averaged with Coot. At your resolution, I would expect to see
the real register
Hi Roger,
I think your ideas are sound, but I would add some "prime-and-switch"
density modification in resolve plus/minus ncs to try and improve the maps
and cut down on phase bias.
Hth,
Dave
--
Hand delivered by Androids
On 1 Sep 2010 16:12, "Roger Rowlett" wrote:
I am trying to find a MR
Hi Qiangmin,
All the comments and references that were already mentioned to you are
excellent,
I would stress 3 points.
1- The detergent.
A clear distinction should be made between the detergent used for
extraction/solubilization and the detergent (or cocktail of
detergents/lipids) used for cryst
POST-DOCTORAL POSITION
DATE OF AVAILABILITY : starting as soon as possible, for 2 years (ANR
contract).
TITLE : purification and crystallization of ABC transporters
POSITION: the position is opened at the Institute for Structural Biology
(IBS) in Grenoble in the team of Jean-Michel Jault and
Thank you for all of your suggestions. I think Pascal Egea has already
summarized the points, and of course Edwin Pozharshi etc. Now I am trying to
confirm if this crystal is detergent or not either using uv microscope or
mass-spec. I also ordered Molecular dimensions stuff. Yes, I noticed that
the
Dear All,
Recently I am working on a protein-DNA complex, and from running
agarose gel, there is a weak delayed band after the band of pure DNA which
indicates some DNA has bind to the protein though the binding efficiency is
low. Then I tried to optimize the condition to increase the bindi
Dear Yang,
You might try acrylamide instead of agarose, since the caging effect of the
tighter matrix might keep the complex together better.
I'm not sure why MgATP? If your protein only binds in the presence of ATP you
might need a non-hydrolyzable analog?
Your concentrations are also vast
Some other things to try:
1. Co-crystallize with a ligand
2. crystallization with lipid cubic phase or bicelles
3. limited proteolysis to define a rigid core
ho
-
Hi all,
I got a crystal of one membrane protein (~60kD) from Na/K phosphate
condition (
Dear All,
My SeMet protein crystals have a needle-like morphology, worse than that of
the native xtals. Also, although the cell dimensions of both forms is very
much similar, the SeMet xtals are twinned(as indicated by Ctruncate plots).
I was wondering if such cases were commonly seen, also is it
Yes, on both counts. SeMet proteins can be frustratingly* close to
their native forms - with little quirks like twinning or lack of
useful diffraction etc. It's common practice to micro (or macro) seed
with native crystals and it often works quite well. It's fun and
sometimes useful to add a few ul
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