Re: [ccp4bb] ligand binding and crystal form

[Artem Evdokimov]

Good (?) afternoon,

I can't resist quoting the facetious answer, once given to me in a similar
situation by the dear departed Felix Frolow: '*proteins are bastards*'.
Well to be exact he used a much less socially acceptable term, and he said
it in Russian - but the point is that sometimes (most of the time?) the
Spider Pig does whatever it wants (yes, this is a Simpsons reference).

What is fascinating is that you have either pure-A situation (and no B) or
pure A+B situation (and again, no pure B). I wonder what would happen if
you were to counter-soak the ligand out. In a related question - how does
the dimer behave kinetically, with respect to site occupancy, Michaelis
number, Kon and Koff? I suspect the answer to your question might have a
lot to do with the cross-talk between two sites in the biologically
relevant dimer coupled with the positional interference in the crystal.
Since you are operating on the time frame similar to the time needed for
molecules to diffuse across a crystal, it is intriguing to think that you
might be experiencing a 'phase locking' (as in Chaos Theory) phenomenon of
some sort with respect to site occupancy - even something as subtle as
crystal orientation might matter (meaning that individual crystals
experience the incoming wave of compound gradient from different crystal
faces which could result in locking of whatever phase that's being
triggered).

So it could be as 'simple' as manifestation of multisite kinetics in a
crystal environment, or as complex as the Butterfy Effect observed in
crystals.

Artem

P.S. I realize that the 'answer' above is more of a series of other
questions, sorry about that.
P.P.S. the cracking of crystals in longer soaks could very well be related
to filling of sites C and D - those are likely empty because when they fill
up their geometry is incompatible with the existing lattice. In a related
case a few years ago I had the same exact situation (i.e. crystals cracked
post-soak) but with some more waiting the cracks 'healed' and the resulting
crystals in fact diffracted - almost as well as the unsoaked ones - and the
space group changed.

- Cosmic Cats approve of this message

www.harkerbio.com
"the rainbow-colored people on our website are not really us"

On Wed, Oct 26, 2016 at 12:47 PM, Veronica Fiorentino <
veronicapfiorent...@gmail.com> wrote:

> Thank you all for responses. I have added my reply below.
>
> On 26 October 2016 at 17:02, Eleanor Dodson 
> wrote:
> > Are your 3 crystals isomorphous, and do they all refine well?
> Yes, all C2221 crystal datasets refine to ~ 19/23 %
> >  I guess you struck lucky with DS3 - as suggested it could be a different
> > soaking, different crystal size, etc, etc..
> >
> > If isomorphous I would compare all 3 structures in COOT and look for
> subtle
> > differences..
> We have.
>
> > Your C222 and C2221 cells are obviously related but have slightly
> different
> > cell volumes so cannot be exactly the same. Do you have any
> > non-crystallographic translation - that can confuse the assignment of 2_1
>
>
> On 26 October 2016 at 16:47, Tristan Croll  wrote:
> > Are these three crystals in order of harvesting (with different soaking
> times)? How big is your ligand. How accessible is the binding pocket (and
> is there a clear difference in accessibility between chains)?
> >
> > T
> >
> Three separate drops; but crystal sizes for C2221 varied 100 -150
> microns. Even the other crystal form C222, was from different drop
> (same crystallisation condition)
> Binding pocket is accessible even in unbound chains. Let me clarify.
> There are 4 chains; 4 binding sites. Two of the binding pockets
> located close to one another; and another 2 are far away from any
> other (even in packing). The sites are close to one another are always
> unoccupied but the sites that are lonely, always have occupied.
>
> I asked this as I wonder if referees might ask _why_ in some chains
> COA is unbound.
>
>
> On 26 October 2016 at 17:11, Zbyszek Otwinowski 
> wrote:
> > What is the height of non-origin Patterson function peak for your data
> sets?
> >
> > have pseudotranslational symmetry that can be detected by calculating the
> > Patterson function. You should have strong reflections with all indexes
> > even or odd, and other reflections being weaker. What is the spot shape
> of
> > these weaker spots?
> >
> > In case of pseudotranslational symmetry, MR can produce a pseudosolution
> > related to the correct one by pseudotranslational symmetry vector.
> > Translate your C2221 solution by {0, 0.5, 0.5} and try refining again.
> >
>
> I ran peakmax within molrep/ccp4i:
>
> List of peaks in order of location  -C222
>  ==
>
>  Peaks related by symmetry are assigned the same site number
>
>  Count Site HeightGrid Fractional coordinates
> Orthogonal coordinates
>
>  WARNING: delta Rho too small for peak determination at 

[ccp4bb] MolMol Question

[Hammond, Robert Glenn]

Hi,


I would like to input all 20 conformers of my ensemble into Molmol and chose 
which of those 20 conformers is the best representation of all of the others. I 
have been looking online for tutorials but I am coming up short. Does anyone 
have any ideas?


Thanks.




Regards,

Robert Hammond

Re: [ccp4bb] ligand binding and crystal form

[Veronica Fiorentino]

Thank you all for responses. I have added my reply below.

On 26 October 2016 at 17:02, Eleanor Dodson  wrote:
> Are your 3 crystals isomorphous, and do they all refine well?
Yes, all C2221 crystal datasets refine to ~ 19/23 %
>  I guess you struck lucky with DS3 - as suggested it could be a different
> soaking, different crystal size, etc, etc..
>
> If isomorphous I would compare all 3 structures in COOT and look for subtle
> differences..
We have.

> Your C222 and C2221 cells are obviously related but have slightly  different
> cell volumes so cannot be exactly the same. Do you have any
> non-crystallographic translation - that can confuse the assignment of 2_1


On 26 October 2016 at 16:47, Tristan Croll  wrote:
> Are these three crystals in order of harvesting (with different soaking 
> times)? How big is your ligand. How accessible is the binding pocket (and is 
> there a clear difference in accessibility between chains)?
>
> T
>
Three separate drops; but crystal sizes for C2221 varied 100 -150
microns. Even the other crystal form C222, was from different drop
(same crystallisation condition)
Binding pocket is accessible even in unbound chains. Let me clarify.
There are 4 chains; 4 binding sites. Two of the binding pockets
located close to one another; and another 2 are far away from any
other (even in packing). The sites are close to one another are always
unoccupied but the sites that are lonely, always have occupied.

I asked this as I wonder if referees might ask _why_ in some chains
COA is unbound.


On 26 October 2016 at 17:11, Zbyszek Otwinowski  wrote:
> What is the height of non-origin Patterson function peak for your data sets?
>
> have pseudotranslational symmetry that can be detected by calculating the
> Patterson function. You should have strong reflections with all indexes
> even or odd, and other reflections being weaker. What is the spot shape of
> these weaker spots?
>
> In case of pseudotranslational symmetry, MR can produce a pseudosolution
> related to the correct one by pseudotranslational symmetry vector.
> Translate your C2221 solution by {0, 0.5, 0.5} and try refining again.
>

I ran peakmax within molrep/ccp4i:

List of peaks in order of location  -C222
 ==

 Peaks related by symmetry are assigned the same site number

 Count Site HeightGrid Fractional coordinates
Orthogonal coordinates

 WARNING: delta Rho too small for peak determination at 263  52   9,
delrho values  0.6313E-03  0.E+00
 WARNING: delta Rho too small for peak determination at 263  60   9,
delrho values  0.6313E-03  0.E+00


*  8000 peaks found, check value of RMN
  NOP highest peaks are selected from these
   11100.00 0   0   0   0.  0.  0.
0.00   0.00   0.00
   22 14.2123   0   0   0.2059  0.  0.
30.27   0.00   0.00
   33 13.6530   0   0   0.2680  0.  0.
39.40   0.00   0.00
   44 14.3951   0   0   0.4548  0.  0.
66.87   0.00   0.00
   55 14.1356   0   0   0.5000  0.  0.
73.53   0.00   0.00
   64 14.3961   0   0   0.5452  0.  0.
80.18   0.00   0.00
   73 13.6582   0   0   0.7320  0.  0.
107.65   0.00   0.00
   82 14.2189   0   0   0.7941  0.  0.
116.78   0.00   0.00
   91 63.04   111   0   0   0.9911  0.  0.
145.74   0.00   0.00
  106 14.3136   1   0   0.3201  0.0025  0.
47.07   0.90   0.00
  116 14.3176   1   0   0.6799  0.0025  0.
99.98   0.90   0.00
  127 13.9341   2   0   0.3670  0.0076  0.
53.96   2.70   0.00
  138 14.2548   2   0   0.4283  0.0093  0.
62.98   3.28   0.00
  148 14.2564   2   0   0.5717  0.0093  0.
84.07   3.28   0.00
  157 13.9371   2   0   0.6330  0.0076  0.
93.09   2.70   0.00
  169 14.0310   3   0   0.0908  0.0129  0.
13.36   4.57   0.00
  17   10 14.7816   3   0   0.1439  0.0096  0.
21.16   3.39   0.00
  18   11 14.1529   3   0   0.2597  0.0129  0.
38.19   4.56   0.00
  19   12 13.8333   3   0   0.2915  0.0111  0.
42.86   3.93   0.00
  20   12 13.8379   3   0   0.7085  0.0111  0.
104.19   3.93   0.00
  21   11 14.1583   3   0   0.7403  0.0129  0.
108.86   4.56   0.00
  22   10 14.7896   3   0   0.8561  0.0096  0.
125.89   3.39   0.00
  239 14.03   102   3   0   0.9092  0.0129  0.
133.69   4.57   0.00
  24   13 14.2225   4   0   0.2259  0.0138  0.
33.22   4.88   0.00
  25   14 15.0256   4   0   0.5000  0.0158  0.
73.53   5.60   0.00
  26   13 14.2287   4   0   0.7741  0.0138  0.
113.83   4.88   0.00
  27   15 13.9519   5   0   0.1734  0.0204 

Re: [ccp4bb] New version SHELXL 2016/6 and PDB2INS

[George Sheldrick]

Dear Kay,

Exactly. That is why we provided two Linux versions (called pdb2ins and 
pdb2ins_oldlinux) on the SHELX server.
So far two versions seem to be enough and for Macs and Windows only one 
version seems to be required. If
anyone finds this to be insufficient, please let Anna know giving 
details of which OS or Linux distribution you

are using.

At the moment we are only providing 64-bit versions of pdb2ins and there 
have been no complaints about that.
This raises the question of whether I am wasting my time providing both 
32- and 64-bit versions of all the SHELX

programs?!

Best wishes, George
CC Anna

On 10/26/2016 05:12 PM, Kay Diederichs wrote:

Hi Phil,

the error message occurs because your glibc is too old for this binary. The 
glibc cannot easily be updated,  it is tied to the distribution, in your case 
RHEL6. A newer kernel does not change this, nor would an update to RHEL 6.8
This is why distributed Linux software should be built on dated distributions.

Best,

Kay

On Wed, 26 Oct 2016 12:11:25 +0100, Phil Evans  wrote:


Hi George

I�m not sure that anyone here is using shelxl but I�ve just updated our general 
64-bit Linux versions anyway. shell starts OK, but with pdb2ins I get a 
complaint. Do you understand this? (I should probably ask our local guys)

Our system is
Scientific Linux release 6.7 (Carbon)

./pdb2ins
./pdb2ins: /lib64/libc.so.6: version `GLIBC_2.14' not found (required by 
/tmp/_MEIQO85tj/libz.so.1)

best wishes
Phil



On 25 Oct 2016, at 20:31, George Sheldrick  wrote:

SHELXL may be used to refine both small and macromolecular structures against 
X-ray or neutron diffraction data, including non-merohedral twins. A new 
version 2016/6 of SHELXL may now be downloaded from the SHELX server. It has 
been well tested by about 20 volunteers to whom I am very grateful. A new 
version of Anna Luebben's program PDB2INS is also available there (for 64 bit 
systems only). PDB2INS makes the preparation of the .ins and .hkl files to run 
macromolecular SHELXL refinements much easier. For most structures deposited in 
the PDB since 2008 these two files can be generated automatically and no 
changes are needed to run SHELXL.

George
--
Prof. George M. Sheldrick FRS
Dept. Structural Chemistry,
University of Goettingen,
Tammannstr. 4,
D37077 Goettingen, Germany
Tel. +49-551-39-33021 or -33068
Fax. +49-551-39-22582





--
Prof. George M. Sheldrick FRS
Dept. Structural Chemistry,
University of Goettingen,
Tammannstr. 4,
D37077 Goettingen, Germany
Tel. +49-551-39-33021 or -33068
Fax. +49-551-39-22582

Re: [ccp4bb] ligand binding and crystal form

[Pierre Rizkallah]

Hello Veronica,

There are 2 aspects to your enquiry:

1- The different number of ligands in your structure could be easily sorted 
out by a longer soak. You don’t have to stick with 1 minute soak. A day or 
longer (I’ve heard of a case in which the soak went on for 30 days), would fill 
up as many sites as can take your ligand. Of course, the soak would have to be 
neutral to the crystal, i.e. won’t cause any damage.

2- C222 and C2221 are interchangeable if you shift the origin by 0.25in the 
x direction. You can try reindexing the mtz file to call the space group C222 
and solve the structure again, and you will see all structures match up. Or you 
can simply shift the solution in C2221 by ¼ in X, and you get the same stats, 
identical. I have done this in the past. But there is one thing you need to do 
first: You need to permute the C2221 axes so that they are 217.4 354.3 147.0. 
But this will change the hand, because it isn’t a cyclic permutation. It is 
like looking down the b-axis with one version or up the b-axis with the other. 
There is a switch in REINDEX to allow to do that. The b-axis is some 9A longer, 
but apart from that there will not be much difference. The two cells are the 
same, effectively. Note that the rogue cell is longer than the other in all 3 
dimensions, around 1 or 2 A in x and z, but 9 in y. If you look in the image 
processing log file, you will see the distance has refined to a slightly 
different value for this data set, provided you have used the same system for 
recording the diffraction patterns. The rogue cell could still have a hidden 
surprise.

3- A proviso for this game of reindexing and origin shift to work, is that 
the shift should be done in the z-direction, depending on how your molecules 
pack in the cell. Symmetry and pseudo-symmetry are fascinating games.

I hope this gives you a different perspective. Good Luck.

Pierre
***
Dr Pierre Rizkallah, Senior Lecturer Structural Biology
Institute of Infection & Immunology, Sir Geraint Evans Building,
School of Medicine, Heath Campus, Cardiff, CF14 4XN
email: rizkall...@cardiff.ac.ukphone: 
+44 29 2074 2248
http://www.cardiff.ac.uk/people/view/126690-rizkallah-pierre

From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Veronica 
Fiorentino
Sent: 26 October 2016 13:32
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] ligand binding and crystal form

Hello all,
I just solved a NCS-tetrameric (biological assembly is just a dimer) crystal 
structures with ligand soak (same plate - same conditions). No density for 
ligand is observed in the first map. In the 2nd, I have 1 ligand bound. In the 
3rd, I have 2 ligands bound. Is there any reason for this 'random' behaviour?
In addition, I observed just one crystal out of 20 gave a different unit cell. 
Pointless confirms to me
"Best Solution:space group C 2 2 2". REFMAC refinement shows R/Rfree ~ 
20/25 %
Cell from mtz :   216.5   345.8   145.290.090.090.0
Space group from mtz: number -   21; name - C 2 2 2

All other datasets have:
Cell from mtz :   147.0   354.3   217.490.090.090.0
Space group from mtz: number -   20; name - C 2 2 21
I tried re-processing/refining the C2221 dataset in C222 but R/Rfree stays 
~45%. Can I also consider the C2221 dataset as a 'different crystal form'?

Am I safe?
Thank you all,
Veronica

Re: [ccp4bb] New version SHELXL 2016/6 and PDB2INS

[R.D. Oeffner]

In line with Kay's message this sounds like pdb2ins having being linked 
statically with glibc.
AFAIK this can be problematic since glibc in reality can never be 
completely statically linked. This may cause version conflicts if the 
same version of glibc is not present on the users system. To avoid this 
problem do not link glibc statically into a binary for distribution.


Correct me if I'm wrong.

Rob






On 26/10/2016 12:11, Phil Evans wrote:

Hi George

I’m not sure that anyone here is using shelxl but I’ve just updated
our general 64-bit Linux versions anyway. shell starts OK, but with
pdb2ins I get a complaint. Do you understand this? (I should probably
ask our local guys)

Our system is
Scientific Linux release 6.7 (Carbon)

 ./pdb2ins
./pdb2ins: /lib64/libc.so.6: version `GLIBC_2.14' not found (required
by /tmp/_MEIQO85tj/libz.so.1)

best wishes
Phil


On 25 Oct 2016, at 20:31, George Sheldrick 
 wrote:


SHELXL may be used to refine both small and macromolecular structures 
against X-ray or neutron diffraction data, including non-merohedral 
twins. A new version 2016/6 of SHELXL may now be downloaded from the 
SHELX server. It has been well tested by about 20 volunteers to whom I 
am very grateful. A new version of Anna Luebben's program PDB2INS is 
also available there (for 64 bit systems only). PDB2INS makes the 
preparation of the .ins and .hkl files to run macromolecular SHELXL 
refinements much easier. For most structures deposited in the PDB 
since 2008 these two files can be generated automatically and no 
changes are needed to run SHELXL.


George
--
Prof. George M. Sheldrick FRS
Dept. Structural Chemistry,
University of Goettingen,
Tammannstr. 4,
D37077 Goettingen, Germany
Tel. +49-551-39-33021 or -33068
Fax. +49-551-39-22582

Re: [ccp4bb] Help with MR result

[Claudia Millán Nebot]

Hi Liuqing,

As Eleanor suggested, if your structure is expected to contain alpha
helices, you might
want to give ARCIMBOLDO_LITE a try, as your resolution is still within the
scope of the method.
In case you have any suspicion of the fold that might be and it contains
beta sheets,
you can try ARCIMBOLDO_BORGES with our precomputed libraries.

Good luck,

Claudia

2016-10-26 15:53 GMT+02:00 SUBSCRIBE CCP4BB Liuqing Chen <519198...@163.com>
:

> Hello!
> I have a crystal diffracted to 2Å, and have no homology structure.  As, i
> can't repeat the crystallization, so i try using Balbe and MorDa program,
> the MorDa give a partial model with Q socre 0.46,  Rwork/Rfree 0.50/0.51.
> it's hard to rebuilt using coot, and when i use Buccaner  or Arp/warp， it's
> built model worse.
> I don't know what to do?
> can anyone give me some suggestions?
> Thanks
> Liuqing Chen
>
>

Re: [ccp4bb] New version SHELXL 2016/6 and PDB2INS

[Kay Diederichs]

Hi Phil,

the error message occurs because your glibc is too old for this binary. The 
glibc cannot easily be updated,  it is tied to the distribution, in your case 
RHEL6. A newer kernel does not change this, nor would an update to RHEL 6.8
This is why distributed Linux software should be built on dated distributions.

Best,

Kay

On Wed, 26 Oct 2016 12:11:25 +0100, Phil Evans  wrote:

>Hi George
>
>I�m not sure that anyone here is using shelxl but I�ve just updated our 
>general 64-bit Linux versions anyway. shell starts OK, but with pdb2ins I get 
>a complaint. Do you understand this? (I should probably ask our local guys)
>
>Our system is
>Scientific Linux release 6.7 (Carbon)
>
> ./pdb2ins 
>./pdb2ins: /lib64/libc.so.6: version `GLIBC_2.14' not found (required by 
>/tmp/_MEIQO85tj/libz.so.1)
>
>best wishes
>Phil
>
>
>> On 25 Oct 2016, at 20:31, George Sheldrick  
>> wrote:
>> 
>> SHELXL may be used to refine both small and macromolecular structures 
>> against X-ray or neutron diffraction data, including non-merohedral twins. A 
>> new version 2016/6 of SHELXL may now be downloaded from the SHELX server. It 
>> has been well tested by about 20 volunteers to whom I am very grateful. A 
>> new version of Anna Luebben's program PDB2INS is also available there (for 
>> 64 bit systems only). PDB2INS makes the preparation of the .ins and .hkl 
>> files to run macromolecular SHELXL refinements much easier. For most 
>> structures deposited in the PDB since 2008 these two files can be generated 
>> automatically and no changes are needed to run SHELXL.
>> 
>> George
>> -- 
>> Prof. George M. Sheldrick FRS
>> Dept. Structural Chemistry, 
>> University of Goettingen,
>> Tammannstr. 4,
>> D37077 Goettingen, Germany
>> Tel. +49-551-39-33021 or -33068
>> Fax. +49-551-39-22582
>> 
>> 

Re: [ccp4bb] ligand binding and crystal form

[Zbyszek Otwinowski]

What is the height of non-origin Patterson function peak for your data sets?

C-centered cells:

  216.5   345.8   145.290.090.090.0

and

  147.0   354.3   217.490.090.090.0

are very different; however, they have common subgroup F222 with similar
unit cell parameters. In F222 one can permute the unit cell axes while
preserving symmetry operators.

For these C centered cells to have approximate F222 subgroup they need to
have pseudotranslational symmetry that can be detected by calculating the 
Patterson function. You should have strong reflections with all indexes
even or odd, and other reflections being weaker. What is the spot shape of
these weaker spots?

In case of pseudotranslational symmetry, MR can produce a pseudosolution
related to the correct one by pseudotranslational symmetry vector.
Translate your C2221 solution by {0, 0.5, 0.5} and try refining again.



Zbyszek Otwinowski




> Dear Veronica,
>
> with 1st, 2nd, 3rd map you mean the density for the same dataset after
> three consecutive cycles of building-refining or three different maps from
> three different crystals?
> If it's the first case, it could be fine, it may mean that at each cycle
> you improve the map so you see signal from the different ligand molecules.
> If it's the second case, well, it could be simply an artifact. Are the
> ligands proximal one to another or bind different sites on the protein?
>
> Best
> V.
>
> 2016-10-26 14:32 GMT+02:00 Veronica Fiorentino <
> veronicapfiorent...@gmail.com>:
>
>> Hello all,
>> I just solved a NCS-tetrameric (biological assembly is just a dimer)
>> crystal structures with ligand soak (same plate - same conditions). No
>> density for ligand is observed in the first map. In the 2nd, I have 1
>> ligand bound. In the 3rd, I have 2 ligands bound. Is there any reason
>> for
>> this 'random' behaviour?
>>
>> In addition, I observed just one crystal out of 20 gave a different unit
>> cell. Pointless confirms to me
>> "Best Solution:space group C 2 2 2". REFMAC refinement shows R/Rfree
>> ~
>> 20/25 %
>> Cell from mtz :   216.5   345.8   145.290.090.090.0
>> Space group from mtz: number -   21; name - C 2 2 2
>>
>> All other datasets have:
>> Cell from mtz :   147.0   354.3   217.490.090.090.0
>> Space group from mtz: number -   20; name - C 2 2 21
>>
>> I tried re-processing/refining the C2221 dataset in C222 but R/Rfree
>> stays
>> ~45%. Can I also consider the C2221 dataset as a 'different crystal
>> form'?
>>
>> Am I safe?
>>
>> Thank you all,
>> Veronica
>>
>
>
>
> --
>
> *Valentina Speranzini, PhD*
> European Molecular Biology Laboratory
> Grenoble Outstation
> 71, avenue des Martyrs, CS 90181
> 38042 Grenoble Cedex 9, France
> Web: http://www.embl.fr
> E-mail: vsperanz...@embl.fr
> Tel: +33 (0)4 76 20 7630
>


Zbyszek Otwinowski
UT Southwestern Medical Center at Dallas
5323 Harry Hines Blvd.
Dallas, TX 75390-8816
Tel. 214-645-6385
Fax. 214-645-6353

Re: [ccp4bb] ligand binding and crystal form

[Eleanor Dodson]

Are your 3 crystals isomorphous, and do they all refine well?
 I guess you struck lucky with DS3 - as suggested it could be a different
soaking, different crystal size, etc, etc..

If isomorphous I would compare all 3 structures in COOT and look for subtle
differences..

Your C222 and C2221 cells are obviously related but have slightly
different cell volumes so cannot be exactly the same. Do you have any
non-crystallographic translation - that can confuse the assignment of 2_1
axes.
Eleanor



On 26 October 2016 at 15:47, Tristan Croll  wrote:

> Are these three crystals in order of harvesting (with different soaking
> times)? How big is your ligand. How accessible is the binding pocket (and
> is there a clear difference in accessibility between chains)?
>
> T
>
>
>
> Tristan Croll
> Research Fellow
> Cambridge Institute for Medical Research
> University of Cambridge CB2 0XY
>
>
>
>
> > On 26 Oct 2016, at 13:32, Veronica Fiorentino <
> veronicapfiorent...@gmail.com> wrote:
> >
> > Hello all,
> > I just solved a NCS-tetrameric (biological assembly is just a dimer)
> crystal structures with ligand soak (same plate - same conditions). No
> density for ligand is observed in the first map. In the 2nd, I have 1
> ligand bound. In the 3rd, I have 2 ligands bound. Is there any reason for
> this 'random' behaviour?
> >
> > In addition, I observed just one crystal out of 20 gave a different unit
> cell. Pointless confirms to me
> > "Best Solution:space group C 2 2 2". REFMAC refinement shows R/Rfree
> ~ 20/25 %
> > Cell from mtz :   216.5   345.8   145.290.090.090.0
> > Space group from mtz: number -   21; name - C 2 2 2
> >
> > All other datasets have:
> > Cell from mtz :   147.0   354.3   217.490.090.090.0
> > Space group from mtz: number -   20; name - C 2 2 21
> >
> > I tried re-processing/refining the C2221 dataset in C222 but R/Rfree
> stays ~45%. Can I also consider the C2221 dataset as a 'different crystal
> form'?
> >
> > Am I safe?
> >
> > Thank you all,
> > Veronica
>

Re: [ccp4bb] ligand binding and crystal form

[Tristan Croll]

Are these three crystals in order of harvesting (with different soaking times)? 
How big is your ligand. How accessible is the binding pocket (and is there a 
clear difference in accessibility between chains)?

T

 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 26 Oct 2016, at 13:32, Veronica Fiorentino  
> wrote:
> 
> Hello all,
> I just solved a NCS-tetrameric (biological assembly is just a dimer) crystal 
> structures with ligand soak (same plate - same conditions). No density for 
> ligand is observed in the first map. In the 2nd, I have 1 ligand bound. In 
> the 3rd, I have 2 ligands bound. Is there any reason for this 'random' 
> behaviour? 
> 
> In addition, I observed just one crystal out of 20 gave a different unit 
> cell. Pointless confirms to me 
> "Best Solution:space group C 2 2 2". REFMAC refinement shows R/Rfree ~ 
> 20/25 %
> Cell from mtz :   216.5   345.8   145.290.090.090.0
> Space group from mtz: number -   21; name - C 2 2 2
> 
> All other datasets have:
> Cell from mtz :   147.0   354.3   217.490.090.090.0
> Space group from mtz: number -   20; name - C 2 2 21
> 
> I tried re-processing/refining the C2221 dataset in C222 but R/Rfree stays 
> ~45%. Can I also consider the C2221 dataset as a 'different crystal form'?
> 
> Am I safe?
> 
> Thank you all,
> Veronica

Re: [ccp4bb] ligand binding and crystal form

[Veronica Fiorentino]

Hi Valentina,

> I just solved a NCS-tetrameric (biological assembly is just a dimer) crystal 
> structures with ligand soak (same plate - same conditions). No density for 
> ligand is observed in the first map. In the 2nd, I have 1 ligand bound. In 
> the 3rd, I have 2 ligands bound. Is there any reason for this 'random' 
> behaviour?

To _clarify_, I have 3 separate crystals; 3 datasets. Soaking time was
approximately 1 minute for all. Same ligand.

DS 1 - > no ligand
DS 2 -> 1 ligand (chain A)
DS 3 -> 2 ligands (chain A & B ; No lignad C).
These three datasets have
Cell from mtz :   147.0   354.3   217.490.090.090.0
Space group from mtz: number -   20; name - C 2 2 21

>
> In addition, I observed just one crystal out of 20 gave a different unit 
> cell. Pointless confirms to me
> "Best Solution:space group C 2 2 2". REFMAC refinement shows R/Rfree ~ 
> 20/25 %
> Cell from mtz :   216.5   345.8   145.290.090.090.0
> Space group from mtz: number -   21; name - C 2 2 2
>
> All other datasets have:
> Cell from mtz :   147.0   354.3   217.490.090.090.0
> Space group from mtz: number -   20; name - C 2 2 21
>
> I tried re-processing/refining the C2221 dataset in C222 but R/Rfree stays 
> ~45%. Can I also consider the C2221 dataset as a 'different crystal form'?
>
> Am I safe?
>
> Thank you all,
> Veronica

Re: [ccp4bb] ligand binding and crystal form

[Valentina Speranzini]

Dear Veronica,

with 1st, 2nd, 3rd map you mean the density for the same dataset after
three consecutive cycles of building-refining or three different maps from
three different crystals?
If it's the first case, it could be fine, it may mean that at each cycle
you improve the map so you see signal from the different ligand molecules.
If it's the second case, well, it could be simply an artifact. Are the
ligands proximal one to another or bind different sites on the protein?

Best
V.

2016-10-26 14:32 GMT+02:00 Veronica Fiorentino <
veronicapfiorent...@gmail.com>:

> Hello all,
> I just solved a NCS-tetrameric (biological assembly is just a dimer)
> crystal structures with ligand soak (same plate - same conditions). No
> density for ligand is observed in the first map. In the 2nd, I have 1
> ligand bound. In the 3rd, I have 2 ligands bound. Is there any reason for
> this 'random' behaviour?
>
> In addition, I observed just one crystal out of 20 gave a different unit
> cell. Pointless confirms to me
> "Best Solution:space group C 2 2 2". REFMAC refinement shows R/Rfree ~
> 20/25 %
> Cell from mtz :   216.5   345.8   145.290.090.090.0
> Space group from mtz: number -   21; name - C 2 2 2
>
> All other datasets have:
> Cell from mtz :   147.0   354.3   217.490.090.090.0
> Space group from mtz: number -   20; name - C 2 2 21
>
> I tried re-processing/refining the C2221 dataset in C222 but R/Rfree stays
> ~45%. Can I also consider the C2221 dataset as a 'different crystal form'?
>
> Am I safe?
>
> Thank you all,
> Veronica
>



-- 

*Valentina Speranzini, PhD*
European Molecular Biology Laboratory
Grenoble Outstation
71, avenue des Martyrs, CS 90181
38042 Grenoble Cedex 9, France
Web: http://www.embl.fr
E-mail: vsperanz...@embl.fr
Tel: +33 (0)4 76 20 7630

Re: [ccp4bb] Help with MR result

[Christian Roth]

Hi,

it seems you haven't found a solution. If there is no model you might 
want to try experimental phasing. With that resolution and the right sg 
a S-SAD might be possible. So you might not even need a HA derivative.


Cheers

Christian



Am 26.10.2016 um 14:53 schrieb SUBSCRIBE CCP4BB Liuqing Chen:

Hello!
I have a crystal diffracted to 2Å, and have no homology structure.  As, i can't 
repeat the crystallization, so i try using Balbe and MorDa program, the MorDa 
give a partial model with Q socre 0.46,  Rwork/Rfree 0.50/0.51.  it's hard to 
rebuilt using coot, and when i use Buccaner  or Arp/warp， it's built model 
worse.
I don't know what to do?
can anyone give me some suggestions?
Thanks
Liuqing Chen

[ccp4bb] Help with MR result

[SUBSCRIBE CCP4BB Liuqing Chen]

Hello!
I have a crystal diffracted to 2Å, and have no homology structure.  As, i can't 
repeat the crystallization, so i try using Balbe and MorDa program, the MorDa 
give a partial model with Q socre 0.46,  Rwork/Rfree 0.50/0.51.  it's hard to 
rebuilt using coot, and when i use Buccaner  or Arp/warp， it's built model 
worse.
I don't know what to do?
can anyone give me some suggestions?
Thanks
Liuqing Chen

Re: [ccp4bb] New version SHELXL 2016/6 and PDB2INS

[Tim Gruene]

Dear Phil,

as far as I understand, SL version 6 builds on kernel 2.6.32, which was 
released nearly a decade ago. Maybe it's time to update? 
Even the rather conservative Debian stable uses 3.16 ...

Best,
Tim

On Wednesday, October 26, 2016 12:11:25 PM Phil Evans wrote:
> Hi George
> 
> I’m not sure that anyone here is using shelxl but I’ve just updated our
> general 64-bit Linux versions anyway. shell starts OK, but with pdb2ins I
> get a complaint. Do you understand this? (I should probably ask our local
> guys)
> 
> Our system is
> Scientific Linux release 6.7 (Carbon)
> 
>  ./pdb2ins
> ./pdb2ins: /lib64/libc.so.6: version `GLIBC_2.14' not found (required by
> /tmp/_MEIQO85tj/libz.so.1)
> 
> best wishes
> Phil
> 
> > On 25 Oct 2016, at 20:31, George Sheldrick 
> > wrote:
> > 
> > SHELXL may be used to refine both small and macromolecular structures
> > against X-ray or neutron diffraction data, including non-merohedral
> > twins. A new version 2016/6 of SHELXL may now be downloaded from the
> > SHELX server. It has been well tested by about 20 volunteers to whom I am
> > very grateful. A new version of Anna Luebben's program PDB2INS is also
> > available there (for 64 bit systems only). PDB2INS makes the preparation
> > of the .ins and .hkl files to run macromolecular SHELXL refinements much
> > easier. For most structures deposited in the PDB since 2008 these two
> > files can be generated automatically and no changes are needed to run
> > SHELXL.
> > 
> > George
> > --
> > Prof. George M. Sheldrick FRS
> > Dept. Structural Chemistry,
> > University of Goettingen,
> > Tammannstr. 4,
> > D37077 Goettingen, Germany
> > Tel. +49-551-39-33021 or -33068
> > Fax. +49-551-39-22582
-- 
--
Paul Scherrer Institut
Dr. Tim Gruene
- persoenlich -
Principal Investigator
Biology and Chemistry
OFLC/102
CH-5232 Villigen PSI

Phone: +41 (0)56 310 5297

GPG Key ID = A46BEE1A



signature.asc
Description: This is a digitally signed message part.

[ccp4bb] ligand binding and crystal form

[Veronica Fiorentino]

Hello all,
I just solved a NCS-tetrameric (biological assembly is just a dimer)
crystal structures with ligand soak (same plate - same conditions). No
density for ligand is observed in the first map. In the 2nd, I have 1
ligand bound. In the 3rd, I have 2 ligands bound. Is there any reason for
this 'random' behaviour?

In addition, I observed just one crystal out of 20 gave a different unit
cell. Pointless confirms to me
"Best Solution:space group C 2 2 2". REFMAC refinement shows R/Rfree ~
20/25 %
Cell from mtz :   216.5   345.8   145.290.090.090.0
Space group from mtz: number -   21; name - C 2 2 2

All other datasets have:
Cell from mtz :   147.0   354.3   217.490.090.090.0
Space group from mtz: number -   20; name - C 2 2 21

I tried re-processing/refining the C2221 dataset in C222 but R/Rfree stays
~45%. Can I also consider the C2221 dataset as a 'different crystal form'?

Am I safe?

Thank you all,
Veronica

Re: [ccp4bb] New version SHELXL 2016/6 and PDB2INS

[Anna Luebben]

Dear Phil,

Thank you for the information.
We made two versions for Linux available;
one packaged on an up-to-date Linux computer and one from an 'older' 
system.
Should there be a library problem with the 'linux' version, the 
'oldllinux' version could work.


best wishes,
Anna


On 10/26/2016 12:11 PM, Phil Evans wrote:

Hi George

I’m not sure that anyone here is using shelxl but I’ve just updated our general 
64-bit Linux versions anyway. shell starts OK, but with pdb2ins I get a 
complaint. Do you understand this? (I should probably ask our local guys)

Our system is
Scientific Linux release 6.7 (Carbon)

  ./pdb2ins
./pdb2ins: /lib64/libc.so.6: version `GLIBC_2.14' not found (required by 
/tmp/_MEIQO85tj/libz.so.1)

best wishes
Phil



On 25 Oct 2016, at 20:31, George Sheldrick  wrote:

SHELXL may be used to refine both small and macromolecular structures against 
X-ray or neutron diffraction data, including non-merohedral twins. A new 
version 2016/6 of SHELXL may now be downloaded from the SHELX server. It has 
been well tested by about 20 volunteers to whom I am very grateful. A new 
version of Anna Luebben's program PDB2INS is also available there (for 64 bit 
systems only). PDB2INS makes the preparation of the .ins and .hkl files to run 
macromolecular SHELXL refinements much easier. For most structures deposited in 
the PDB since 2008 these two files can be generated automatically and no 
changes are needed to run SHELXL.

George
--
Prof. George M. Sheldrick FRS
Dept. Structural Chemistry,
University of Goettingen,
Tammannstr. 4,
D37077 Goettingen, Germany
Tel. +49-551-39-33021 or -33068
Fax. +49-551-39-22582

Re: [ccp4bb] New version SHELXL 2016/6 and PDB2INS

[Martín Martínez Ripoll]

Mac version runs OK!

Martin
_
Dr. Martin Martinez-Ripoll
Research Professor
xmar...@iqfr.csic.es
Department of Crystallography & Structural Biology
www.xtal.iqfr.csic.es
www.xtal.iqfr.csic.es/Cristalografia/ 
Telf.: +34 917459550
Consejo Superior de Investigaciones Científicas
Spanish National Research Council


-Mensaje original-
De: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] En nombre de Phil
Evans
Enviado el: miércoles, 26 de octubre de 2016 13:11
Para: CCP4BB@JISCMAIL.AC.UK
Asunto: Re: [ccp4bb] New version SHELXL 2016/6 and PDB2INS

Hi George

I’m not sure that anyone here is using shelxl but I’ve just updated our
general 64-bit Linux versions anyway. shell starts OK, but with pdb2ins I
get a complaint. Do you understand this? (I should probably ask our local
guys)

Our system is
Scientific Linux release 6.7 (Carbon)

 ./pdb2ins 
./pdb2ins: /lib64/libc.so.6: version `GLIBC_2.14' not found (required by
/tmp/_MEIQO85tj/libz.so.1)

best wishes
Phil


> On 25 Oct 2016, at 20:31, George Sheldrick 
wrote:
> 
> SHELXL may be used to refine both small and macromolecular structures
against X-ray or neutron diffraction data, including non-merohedral twins. A
new version 2016/6 of SHELXL may now be downloaded from the SHELX server. It
has been well tested by about 20 volunteers to whom I am very grateful. A
new version of Anna Luebben's program PDB2INS is also available there (for
64 bit systems only). PDB2INS makes the preparation of the .ins and .hkl
files to run macromolecular SHELXL refinements much easier. For most
structures deposited in the PDB since 2008 these two files can be generated
automatically and no changes are needed to run SHELXL.
> 
> George
> -- 
> Prof. George M. Sheldrick FRS
> Dept. Structural Chemistry, 
> University of Goettingen,
> Tammannstr. 4,
> D37077 Goettingen, Germany
> Tel. +49-551-39-33021 or -33068
> Fax. +49-551-39-22582
> 
> 

Re: [ccp4bb] New version SHELXL 2016/6 and PDB2INS

[Phil Evans]

Hi George

I’m not sure that anyone here is using shelxl but I’ve just updated our general 
64-bit Linux versions anyway. shell starts OK, but with pdb2ins I get a 
complaint. Do you understand this? (I should probably ask our local guys)

Our system is
Scientific Linux release 6.7 (Carbon)

 ./pdb2ins 
./pdb2ins: /lib64/libc.so.6: version `GLIBC_2.14' not found (required by 
/tmp/_MEIQO85tj/libz.so.1)

best wishes
Phil


> On 25 Oct 2016, at 20:31, George Sheldrick  
> wrote:
> 
> SHELXL may be used to refine both small and macromolecular structures against 
> X-ray or neutron diffraction data, including non-merohedral twins. A new 
> version 2016/6 of SHELXL may now be downloaded from the SHELX server. It has 
> been well tested by about 20 volunteers to whom I am very grateful. A new 
> version of Anna Luebben's program PDB2INS is also available there (for 64 bit 
> systems only). PDB2INS makes the preparation of the .ins and .hkl files to 
> run macromolecular SHELXL refinements much easier. For most structures 
> deposited in the PDB since 2008 these two files can be generated 
> automatically and no changes are needed to run SHELXL.
> 
> George
> -- 
> Prof. George M. Sheldrick FRS
> Dept. Structural Chemistry, 
> University of Goettingen,
> Tammannstr. 4,
> D37077 Goettingen, Germany
> Tel. +49-551-39-33021 or -33068
> Fax. +49-551-39-22582
> 
>