Re: [ccp4bb] structure with missing density

[Ditlev Egeskov Brodersen]

Dear Deepanshu,

Is the primary sequence in the neighbourhood of your missing strands conserved? 
Perhaps that part differs in your structure relative to the search model. Have 
you tried omitting the region from MR and rerunning refinement without to see 
if some new density pops up?

If not, although this doesn’t sound like a classical flexible region, it may 
still be disordered. If you really can’t build it, you can of course leave it 
out of the structure and deposit anyway. The rest of the structure might be 
very relevant for other people.

I also note that your refinement R factor is a bit low relative to the Rfree, 
which is indicating overfitting. Do you use individual B factors? You probably 
should use grouped Bs at 3 Å.

Best,

Ditlev

---

Ditlev E. Brodersen
Lektor, Associate Professor
Department of Molecular Biology and Genetics, Aarhus University
Gustav Wieds Vej 10c, DK-8000 Aarhus C, Denmark

Phone:  +45 21669001
Lab home page: www.bioxray.au.dk/~deb
Educational IT portal: curriculearn.dk

Sent from my iPhone

Den 10. okt. 2018 kl. 04.59 skrev Deepanshu Choudhary 
mailto:deepanshui...@gmail.com>>:

Dear all,
I am working on a protein complex. After many efforts, I obtained crystals from 
one of the refinement screen (which took few weeks to grow) and I got a 3 Å 
dataset at synchrotron. I scaled the dataset in P21 spacegroup (which is also 
confirmed by Zanuda and Pointless). There is no twinning detected. I solved the 
phase using molecular replacement with a model of over 90% sequence identity. 
After several rounds of refinement with Refmac, the Rfree is 0.307 and Rfactor 
of 0.23.
The density looks good and I can see everything that's important. But one of 
the proteins has missing density in 2 of its beta strands (corresponding to 
~15%) and its not appearing upon several rounds of refinement. Also, the B 
factors are higher for this protein. The missing beta strands are not at the 
interface of the complex.
I ran some crystals on the gel and did silver staining to find both the 
proteins and no degradation products. I doubt that there is any proteolytic 
cleavage because the protein is unlikely to remain folded if those beta strands 
are chopped out.
I want to ask if such a structure with missing density and high B-factors 
(>100) can be deposited. Is it possible that some parts of the lattice don't 
have this protein with missing density which is resulting in high B factors?
I would appreciate your efforts if someone can send me few references 
describing such type of structures. I would also welcome any other suggestions 
and recommendations.

Thanks and regards,
Deepanshu



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

[ccp4bb] structure with missing density

[Deepanshu Choudhary]

Dear all,
I am working on a protein complex. After many efforts, I obtained crystals
from one of the refinement screen (which took few weeks to grow) and I got
a 3 Å dataset at synchrotron. I scaled the dataset in P21 spacegroup (which
is also confirmed by Zanuda and Pointless). There is no twinning detected.
I solved the phase using molecular replacement with a model of over 90%
sequence identity. After several rounds of refinement with Refmac, the
Rfree is 0.307 and Rfactor of 0.23.
The density looks good and I can see everything that's important. But one
of the proteins has missing density in 2 of its beta strands (corresponding
to ~15%) and its not appearing upon several rounds of refinement. Also, the
B factors are higher for this protein. The missing beta strands are not at
the interface of the complex.
I ran some crystals on the gel and did silver staining to find both the
proteins and no degradation products. I doubt that there is any proteolytic
cleavage because the protein is unlikely to remain folded if those beta
strands are chopped out.
I want to ask if such a structure with missing density and high B-factors
(>100) can be deposited. Is it possible that some parts of the lattice
don't have this protein with missing density which is resulting in high B
factors?
I would appreciate your efforts if someone can send me few references
describing such type of structures. I would also welcome any other
suggestions and recommendations.

Thanks and regards,
Deepanshu



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

Re: [ccp4bb] Issue with high Rfree (0.25) for a high-resolution dataset (1.05 Ang)

[Eleanor Dodson]

I do think at times the Rfree is a bit of a sacred cow . At one level if
the maps look OK not much can be wrong..
Is the Ramachandran plot OK?

You dont say what the R factor is but does this mean there is a large
difference between the Rfactor and Rfree?

If you are using REFMAC look at the plots of  R factor and Rfree v
resolution? Is there a big divergence at some points? Could you have
included overloads? Are there icerings?

Then look at the plot of  v  v resolution/
Is the scaling procedure meaning these diverge in some regions?
Is there some strange effects at low resolution?

And so on and so on..
Eleanor



On Tue, 9 Oct 2018 at 19:31, Wim Burmeister  wrote:

> Hello,
> at that resolution, the refinement of anisotropic atomic B-factors is
> absolutely required, as is the modelisation of alternate conformations for
> surface residues. Optimize also the weight of different restraints (for
> exemple on B-factors) in order to get the lowest Rfree.
> Best
> Wim
>
> --
> *De: *"Robbie Joosten" 
> *À: *CCP4BB@JISCMAIL.AC.UK
> *Envoyé: *Mardi 9 Octobre 2018 20:02:17
> *Objet: *Re: [ccp4bb] Issue with high Rfree (0.25) for a high-resolution
> dataset (1.05 Ang)
>
> Hi Hugo,
>
> Perhaps you should play with your refinement strategy. Use a decent number
> of cycles and a sensible restraint weight (something that gives you rmsZ <
> 1.0 and good R-factors). Anisotropic B-factors are probably needed and make
> your model as complete as your maps allow.
>
> You could try pdb-redo to see if this can help you on your way.
>
> Cheers,
> Robbie
>
> On 9 Oct 2018 19:12, Guto Rhys  wrote:
>
> Hi all,
>
> I have a 1.05 Angstrom dataset that I was able to phase but the refined
> model only has an Rfree of approximately 0.25. The dataset includes 1800
> images and, as the crystal did not suffer significantly from radiation
> damage, comprises all 360 deg. Auto-processing pipelines at diamond light
> source all suggest I222. I have also indexed the data in iMOSFLM, which has
> the highest-symmetry Laue group that is least penalised of I422. Subsequent
> scaling and merging in AIMLESS strongly indicates that I222 is the likely
> space group (see below). I have ran the refined model through ZANUDA, which
> has similar R values to lower symmetry space groups (see below). The output
> from Phenix Xtriage does not find any specific crystal pathologies and if
> twinning is present it is very low (2 to 4%, see below). The difference map
> suggests that the model accounts for nearly all the density. Any ideas or
> direction would be greatly appreciated.
>
> Best,
> Guto
>
>
> AIMLESS Summary
>Overall  InnerShell  OuterShell
> Low resolution limit   27.75 27.75  1.07
> High resolution limit   1.05  5.75  1.05
>
> Rmerge  (within I+/I-) 0.050 0.078 0.466
> Rmerge  (all I+ and I-)0.051 0.080 0.536
> Rmeas (within I+/I-)   0.055 0.086 0.591
> Rmeas (all I+ & I-)0.054 0.085 0.613
> Rpim (within I+/I-)0.023 0.034 0.359
> Rpim (all I+ & I-) 0.017 0.028 0.288
> Rmerge in top intensity bin0.049- -
> Total number of observations  107950   779  1972
> Total number unique1131588   486
> Mean((I)/sd(I)) 19.7  46.2   1.8
> Mn(I) half-set correlation CC(1/2) 0.998 0.994 0.796
> Completeness99.1  99.2  90.4
> Multiplicity 9.5   8.9   4.1
>
> Anomalous completeness  98.1 100.0  79.1
> Anomalous multiplicity   5.0   6.4   2.2
> DelAnom correlation between half-sets -0.067 0.286 0.097
> Mid-Slope of Anom Normal Probability   0.789   - -
>
> Estimate of maximum resolution for significant anomalous signal =  1.14A,
> from CCanom >  0.15
>
> Estimates of resolution limits: overall
>from half-dataset correlation CC(1/2) >  0.30: limit =  1.05A  ==
> maximum resolution
>from Mn(I/sd) >  1.50: limit =  1.05A  ==
> maximum resolution
>from Mn(I/sd) >  2.00: limit =  1.07A
>
> Estimates of resolution limits in reciprocal lattice directions:
>   Along h axis
>from half-dataset correlation CC(1/2) >  0.30: limit =  1.06A
>from Mn(I/sd) >  1.50: limit =  1.09A
>   Along k axis
>from half-dataset correlation CC(1/2) >  0.30: limit =  1.11A
>from Mn(I/sd) >  1.50: limit =  1.13A
>   Along l axis
>from half-dataset correlation CC(1/2) >  0.30: limit =  1.05A  ==
> maximum resolution
>from Mn(I/sd) >  1.50: limit =  1.05A
>
> Anisotropic deltaB 

[ccp4bb] CCN resend

[Nigel Moriarty]

Folks

It seems that 50% of people got an email that was 50% greek. I have tried
to remove this by removing formatting. Hope this helps.

The second 2018 issue of the CCN is now available at

http://phenix-online.org/newsletter/

Articles include

·   Fitting tips #16 – Vicinal disulfides: Have you seen one of these
strange gems?

·   CaBLAM: A C-Alpha Based Low-resolution Annotation Method for secondary
structure and validation

·   Tools for interpreting cryo-EM maps using models from the PDB

·   Using the New Program Template62

Cheers

Nigel

---
Nigel W. Moriarty
Building 33R0349, Molecular Biophysics and Integrated Bioimaging
Lawrence Berkeley National Laboratory
Berkeley, CA 94720-8235
Phone : 510-486-5709 Email : nwmoria...@lbl.gov
Fax   : 510-486-5909   Web  : CCI.LBL.gov



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

Re: [ccp4bb] Issue with high Rfree (0.25) for a high-resolution dataset (1.05 Ang)

[Wim Burmeister]

Hello, 
at that resolution, the refinement of anisotropic atomic B-factors is 
absolutely required, as is the modelisation of alternate conformations for 
surface residues. Optimize also the weight of different restraints (for exemple 
on B-factors) in order to get the lowest Rfree. 
Best 
Wim 


De: "Robbie Joosten"  
À: CCP4BB@JISCMAIL.AC.UK 
Envoyé: Mardi 9 Octobre 2018 20:02:17 
Objet: Re: [ccp4bb] Issue with high Rfree (0.25) for a high-resolution dataset 
(1.05 Ang) 

Hi Hugo, 

Perhaps you should play with your refinement strategy. Use a decent number of 
cycles and a sensible restraint weight (something that gives you rmsZ < 1.0 and 
good R-factors). Anisotropic B-factors are probably needed and make your model 
as complete as your maps allow. 

You could try pdb-redo to see if this can help you on your way. 

Cheers, 
Robbie 

On 9 Oct 2018 19:12, Guto Rhys  wrote: 




Hi all, 

I have a 1.05 Angstrom dataset that I was able to phase but the refined model 
only has an Rfree of approximately 0.25. The dataset includes 1800 images and, 
as the crystal did not suffer significantly from radiation damage, comprises 
all 360 deg. Auto-processing pipelines at diamond light source all suggest 
I222. I have also indexed the data in iMOSFLM, which has the highest-symmetry 
Laue group that is least penalised of I422. Subsequent scaling and merging in 
AIMLESS strongly indicates that I222 is the likely space group (see below). I 
have ran the refined model through ZANUDA, which has similar R values to lower 
symmetry space groups (see below). The output from Phenix Xtriage does not find 
any specific crystal pathologies and if twinning is present it is very low (2 
to 4%, see below). The difference map suggests that the model accounts for 
nearly all the density. Any ideas or direction would be greatly appreciated. 

Best, 
Guto 


AIMLESS Summary 
Overall InnerShell OuterShell 
Low resolution limit 27.75 27.75 1.07 
High resolution limit 1.05 5.75 1.05 

Rmerge (within I+/I-) 0.050 0.078 0.466 
Rmerge (all I+ and I-) 0.051 0.080 0.536 
Rmeas (within I+/I-) 0.055 0.086 0.591 
Rmeas (all I+ & I-) 0.054 0.085 0.613 
Rpim (within I+/I-) 0.023 0.034 0.359 
Rpim (all I+ & I-) 0.017 0.028 0.288 
Rmerge in top intensity bin 0.049 - - 
Total number of observations 107950 779 1972 
Total number unique 11315 88 486 
Mean((I)/sd(I)) 19.7 46.2 1.8 
Mn(I) half-set correlation CC(1/2) 0.998 0.994 0.796 
Completeness 99.1 99.2 90.4 
Multiplicity 9.5 8.9 4.1 

Anomalous completeness 98.1 100.0 79.1 
Anomalous multiplicity 5.0 6.4 2.2 
DelAnom correlation between half-sets -0.067 0.286 0.097 
Mid-Slope of Anom Normal Probability 0.789 - - 

Estimate of maximum resolution for significant anomalous signal = 1.14A, from 
CCanom > 0.15 

Estimates of resolution limits: overall 
from half-dataset correlation CC(1/2) > 0.30: limit = 1.05A == maximum 
resolution 
from Mn(I/sd) > 1.50: limit = 1.05A == maximum resolution 
from Mn(I/sd) > 2.00: limit = 1.07A 

Estimates of resolution limits in reciprocal lattice directions: 
Along h axis 
from half-dataset correlation CC(1/2) > 0.30: limit = 1.06A 
from Mn(I/sd) > 1.50: limit = 1.09A 
Along k axis 
from half-dataset correlation CC(1/2) > 0.30: limit = 1.11A 
from Mn(I/sd) > 1.50: limit = 1.13A 
Along l axis 
from half-dataset correlation CC(1/2) > 0.30: limit = 1.05A == maximum 
resolution 
from Mn(I/sd) > 1.50: limit = 1.05A 

Anisotropic deltaB (i.e. range of principal components), A^2: 6.40 

Average unit cell: 29.12 29.26 55.50 90.00 90.00 90.00 
Space group: I 2 2 2 
Average mosaicity: 0.36 

AIMLESS Laue Group prediction 

Laue Group Lklhd NetZc Zc+ Zc- CC CC- Rmeas R- Delta ReindexOperator 

= 1 I m m m *** 0.987 6.25 9.19 2.94 0.92 0.29 0.07 0.49 0.0 [h,k,l] 
2 I 1 2/m 1 0.004 4.36 9.32 4.96 0.93 0.50 0.07 0.30 0.0 [-h,-k,l] 
3 I 1 2/m 1 0.004 4.14 9.24 5.10 0.92 0.51 0.07 0.30 0.0 [k,-h,l] 
4 I 1 2/m 1 0.004 4.05 9.23 5.18 0.92 0.52 0.06 0.31 0.0 [h,-l,k] 
5 I 4/m m m 0.000 6.35 6.35 0.00 0.64 0.00 0.22 0.00 0.3 [h,k,l] 
6 I 4/m 0.000 0.90 6.83 5.93 0.68 0.59 0.17 0.26 0.3 [h,k,l] 
7 P -1 0.000 3.69 9.36 5.67 0.94 0.57 0.06 0.26 0.0 [-h,k,1/2h-1/2k-1/2l] 
8 I 1 2/m 1 0.000 0.08 6.40 6.33 0.64 0.63 0.23 0.22 0.3 
[-1/2h+1/2k-1/2l,-h-k,-1/2h+1/2k+1/2l] 
9 F m m m 0.000 -0.43 6.17 6.60 0.62 0.66 0.24 0.20 0.3 [h+k,-h+k,l] 
10 I 1 2/m 1 0.000 0.75 6.89 6.14 0.69 0.61 0.22 0.22 0.3 
[-1/2h-1/2k-1/2l,h-k,-1/2h-1/2k+1/2l] 


Xtriage Summary 

Twinning and intensity statistics summary (acentric data): 

Statistics independent of twin laws 
/^2 : 2.126 (untwinned: 2.0, perfect twin: 1.5) 
^2/ : 0.774 (untwinned: 0.785, perfect twin: 0.885) 
<|E^2-1|> : 0.761 (untwinned: 0.736, perfect twin: 0.541) 
<|L|>, : 0.490, 0.323 
Multivariate Z score L-test: 1.303 

The multivariate Z score is a quality measure of the given 
spread in intensities. Good to reasonable data are expected 
to have a Z score lower than 3.5. 
Large values can indicate twinning, but small values do not 
necessarily 

Re: [ccp4bb] Issue with high Rfree (0.25) for a high-resolution dataset (1.05 Ang)

[Isupov, Michail]

Hi Guto,

I would be very cautious with the InnerShell merging statistics as yours.
Rmeas in this shell is higher than Rmeas overall,
which probably means you have a lot of overloads,
(or problems with backstop shadow, which is not so likely
on Diamond nowdays).

 And generally InnerShell Rmerge over 5-6% in XDS processing
 (Diamond pipelines) is usually an indicator of future problems in 
refinement.

Rfree of 25 % at 1.06  A seems to confirm your space group. 
In the case of  wrong space group or origin (ZANUDA), it would normally be well 
over 35 %.

Try to collect data  setting transmission/exposure time to lower values.
 If this was the only crystal ever explain high-ish value of freeR in the 
methods
session as due to overexposure.

For cases like yours I would really like to see Rmeas in the inner shell in 
Table 1.

Best wishes,

Misha Isupov


From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Guto Rhys 
[guto.r...@bristol.ac.uk]
Sent: Tuesday, October 9, 2018 6:12 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] Issue with high Rfree (0.25) for a high-resolution dataset 
(1.05 Ang)

Hi all,

I have a 1.05 Angstrom dataset that I was able to phase but the refined model 
only has an Rfree of approximately 0.25. The dataset includes 1800 images and, 
as the crystal did not suffer significantly from radiation damage, comprises 
all 360 deg. Auto-processing pipelines at diamond light source all suggest 
I222. I have also indexed the data in iMOSFLM, which has the highest-symmetry 
Laue group that is least penalised of I422. Subsequent scaling and merging in 
AIMLESS strongly indicates that I222 is the likely space group (see below). I 
have ran the refined model through ZANUDA, which has similar R values to lower 
symmetry space groups (see below). The output from Phenix Xtriage does not find 
any specific crystal pathologies and if twinning is present it is very low (2 
to 4%, see below). The difference map suggests that the model accounts for 
nearly all the density. Any ideas or direction would be greatly appreciated.

Best,
Guto


AIMLESS Summary
   Overall  InnerShell  OuterShell
Low resolution limit   27.75 27.75  1.07
High resolution limit   1.05  5.75  1.05

Rmerge  (within I+/I-) 0.050 0.078 0.466
Rmerge  (all I+ and I-)0.051 0.080 0.536
Rmeas (within I+/I-)   0.055 0.086 0.591
Rmeas (all I+ & I-)0.054 0.085 0.613
Rpim (within I+/I-)0.023 0.034 0.359
Rpim (all I+ & I-) 0.017 0.028 0.288
Rmerge in top intensity bin0.049- -
Total number of observations  107950   779  1972
Total number unique1131588   486
Mean((I)/sd(I)) 19.7  46.2   1.8
Mn(I) half-set correlation CC(1/2) 0.998 0.994 0.796
Completeness99.1  99.2  90.4
Multiplicity 9.5   8.9   4.1

Anomalous completeness  98.1 100.0  79.1
Anomalous multiplicity   5.0   6.4   2.2
DelAnom correlation between half-sets -0.067 0.286 0.097
Mid-Slope of Anom Normal Probability   0.789   - -

Estimate of maximum resolution for significant anomalous signal =  1.14A, from 
CCanom >  0.15

Estimates of resolution limits: overall
   from half-dataset correlation CC(1/2) >  0.30: limit =  1.05A  == maximum 
resolution
   from Mn(I/sd) >  1.50: limit =  1.05A  == maximum 
resolution
   from Mn(I/sd) >  2.00: limit =  1.07A

Estimates of resolution limits in reciprocal lattice directions:
  Along h axis
   from half-dataset correlation CC(1/2) >  0.30: limit =  1.06A
   from Mn(I/sd) >  1.50: limit =  1.09A
  Along k axis
   from half-dataset correlation CC(1/2) >  0.30: limit =  1.11A
   from Mn(I/sd) >  1.50: limit =  1.13A
  Along l axis
   from half-dataset correlation CC(1/2) >  0.30: limit =  1.05A  == maximum 
resolution
   from Mn(I/sd) >  1.50: limit =  1.05A

Anisotropic deltaB (i.e. range of principal components), A^2:  6.40

Average unit cell:   29.12   29.26   55.50   90.00   90.00   90.00
Space group: I 2 2 2
Average mosaicity:   0.36

AIMLESS Laue Group prediction

   Laue GroupLklhd   NetZc  Zc+   Zc-CCCC-  Rmeas   R-  Delta 
ReindexOperator

= 1I m m m  ***  0.987   6.25  9.19  2.94   0.92  0.29   0.07  0.49   0.0 
[h,k,l]
  2  I 1 2/m 1   0.004   4.36  9.32  4.96   0.93  0.50   0.07  0.30   0.0 
[-h,-k,l]
  3  I 1 2/m 1   0.004   4.14  9.24  5.10   0.92  0.51   0.07  0.30   0.0 

Re: [ccp4bb] Issue with high Rfree (0.25) for a high-resolution dataset (1.05 Ang)

[Robbie Joosten]

Hi Hugo,

Perhaps you should play with your refinement strategy. Use a decent number of 
cycles and a sensible restraint weight (something that gives you rmsZ < 1.0 and 
good R-factors). Anisotropic B-factors are probably needed and make your model 
as complete as your maps allow.

You could try pdb-redo to see if this can help you on your way.

Cheers,
Robbie

On 9 Oct 2018 19:12, Guto Rhys  wrote:

Hi all,

I have a 1.05 Angstrom dataset that I was able to phase but the refined model 
only has an Rfree of approximately 0.25. The dataset includes 1800 images and, 
as the crystal did not suffer significantly from radiation damage, comprises 
all 360 deg. Auto-processing pipelines at diamond light source all suggest 
I222. I have also indexed the data in iMOSFLM, which has the highest-symmetry 
Laue group that is least penalised of I422. Subsequent scaling and merging in 
AIMLESS strongly indicates that I222 is the likely space group (see below). I 
have ran the refined model through ZANUDA, which has similar R values to lower 
symmetry space groups (see below). The output from Phenix Xtriage does not find 
any specific crystal pathologies and if twinning is present it is very low (2 
to 4%, see below). The difference map suggests that the model accounts for 
nearly all the density. Any ideas or direction would be greatly appreciated.

Best,
Guto


AIMLESS Summary
   Overall  InnerShell  OuterShell
Low resolution limit   27.75 27.75  1.07
High resolution limit   1.05  5.75  1.05

Rmerge  (within I+/I-) 0.050 0.078 0.466
Rmerge  (all I+ and I-)0.051 0.080 0.536
Rmeas (within I+/I-)   0.055 0.086 0.591
Rmeas (all I+ & I-)0.054 0.085 0.613
Rpim (within I+/I-)0.023 0.034 0.359
Rpim (all I+ & I-) 0.017 0.028 0.288
Rmerge in top intensity bin0.049- -
Total number of observations  107950   779  1972
Total number unique1131588   486
Mean((I)/sd(I)) 19.7  46.2   1.8
Mn(I) half-set correlation CC(1/2) 0.998 0.994 0.796
Completeness99.1  99.2  90.4
Multiplicity 9.5   8.9   4.1

Anomalous completeness  98.1 100.0  79.1
Anomalous multiplicity   5.0   6.4   2.2
DelAnom correlation between half-sets -0.067 0.286 0.097
Mid-Slope of Anom Normal Probability   0.789   - -

Estimate of maximum resolution for significant anomalous signal =  1.14A, from 
CCanom >  0.15

Estimates of resolution limits: overall
   from half-dataset correlation CC(1/2) >  0.30: limit =  1.05A  == maximum 
resolution
   from Mn(I/sd) >  1.50: limit =  1.05A  == maximum 
resolution
   from Mn(I/sd) >  2.00: limit =  1.07A

Estimates of resolution limits in reciprocal lattice directions:
  Along h axis
   from half-dataset correlation CC(1/2) >  0.30: limit =  1.06A
   from Mn(I/sd) >  1.50: limit =  1.09A
  Along k axis
   from half-dataset correlation CC(1/2) >  0.30: limit =  1.11A
   from Mn(I/sd) >  1.50: limit =  1.13A
  Along l axis
   from half-dataset correlation CC(1/2) >  0.30: limit =  1.05A  == maximum 
resolution
   from Mn(I/sd) >  1.50: limit =  1.05A

Anisotropic deltaB (i.e. range of principal components), A^2:  6.40

Average unit cell:   29.12   29.26   55.50   90.00   90.00   90.00
Space group: I 2 2 2
Average mosaicity:   0.36

AIMLESS Laue Group prediction

   Laue GroupLklhd   NetZc  Zc+   Zc-CCCC-  Rmeas   R-  Delta 
ReindexOperator

= 1I m m m  ***  0.987   6.25  9.19  2.94   0.92  0.29   0.07  0.49   0.0 
[h,k,l]
  2  I 1 2/m 1   0.004   4.36  9.32  4.96   0.93  0.50   0.07  0.30   0.0 
[-h,-k,l]
  3  I 1 2/m 1   0.004   4.14  9.24  5.10   0.92  0.51   0.07  0.30   0.0 
[k,-h,l]
  4  I 1 2/m 1   0.004   4.05  9.23  5.18   0.92  0.52   0.06  0.31   0.0 
[h,-l,k]
  5  I 4/m m m   0.000   6.35  6.35  0.00   0.64  0.00   0.22  0.00   0.3 
[h,k,l]
  6  I 4/m   0.000   0.90  6.83  5.93   0.68  0.59   0.17  0.26   0.3 
[h,k,l]
  7   P -1   0.000   3.69  9.36  5.67   0.94  0.57   0.06  0.26   0.0 
[-h,k,1/2h-1/2k-1/2l]
  8  I 1 2/m 1   0.000   0.08  6.40  6.33   0.64  0.63   0.23  0.22   0.3 
[-1/2h+1/2k-1/2l,-h-k,-1/2h+1/2k+1/2l]
  9F m m m   0.000  -0.43  6.17  6.60   0.62  0.66   0.24  0.20   0.3 
[h+k,-h+k,l]
10  I 1 2/m 1   0.000   0.75  6.89  6.14   0.69  0.61   0.22  0.22   0.3 
[-1/2h-1/2k-1/2l,h-k,-1/2h-1/2k+1/2l]


Xtriage Summary

Twinning and intensity statistics summary (acentric data):

Statistics 

[ccp4bb] Computational Crystallography Newsletter

[Nigel Moriarty]

Folks

The second 2018 issue of the CCN is now available at

http://phenix-online.org/newsletter/

Articles include

·   Fitting tips #16 – Vicinal disulfides: Have you seen one of these
strange gems?

·   CaBLAM: A C-Alpha Based Low-resolution Annotation Method for secondary
structure and validation

·   Tools for interpreting cryo-EM maps using models from the PDB

·   Using the New Program Template62

Cheers

Nigel

---
Nigel W. Moriarty
Building 33R0349, Molecular Biophysics and Integrated Bioimaging
Lawrence Berkeley National Laboratory
Berkeley, CA 94720-8235
Phone : 510-486-5709 Email : nwmoria...@lbl.gov
Fax   : 510-486-5909   Web  : CCI.LBL.gov



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

[ccp4bb] Postdoc opening at Ailong Ke's lab, Cornell University

[Ailong Ke]

A NIH-funded postdoctoral position in structural biology is open in Prof. 
Ailong Ke’s laboratory at Cornell 
University, Ithaca, NY, USA. The research projects seek to understand the 
structure and function of RNA-protein complexes in CRISPR-Cas and other 
RNA-guided systems. For representative publications, please refer to: Science. 
2018 Jul 6;361(6397); Nature. 
2017 Oct 5;550(7674):137-141; 
Cell. 2017 Jun 29;170(1):48-60; 
Nature. 2016 Feb 
25;530(7591):499-503.

Applicants are expected to hold (or expect) a Ph.D. or equivalent degree in 
structural biology (crystallography or EM). Prior experiences in  nucleic acid 
biochemistry and/or single molecule techniques are appreciated. Additional 
information can be found at http://sites.google.com/site/kelaboratory/

To apply, please send CV and names of 3 references to: 
ak...@cornell.edu

Cornell University is an equal opportunity, affirmative action educator and 
employer.




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

[ccp4bb] Issue with high Rfree (0.25) for a high-resolution dataset (1.05 Ang)

[Guto Rhys]

Hi all,

I have a 1.05 Angstrom dataset that I was able to phase but the refined model 
only has an Rfree of approximately 0.25. The dataset includes 1800 images and, 
as the crystal did not suffer significantly from radiation damage, comprises 
all 360 deg. Auto-processing pipelines at diamond light source all suggest 
I222. I have also indexed the data in iMOSFLM, which has the highest-symmetry 
Laue group that is least penalised of I422. Subsequent scaling and merging in 
AIMLESS strongly indicates that I222 is the likely space group (see below). I 
have ran the refined model through ZANUDA, which has similar R values to lower 
symmetry space groups (see below). The output from Phenix Xtriage does not find 
any specific crystal pathologies and if twinning is present it is very low (2 
to 4%, see below). The difference map suggests that the model accounts for 
nearly all the density. Any ideas or direction would be greatly appreciated.

Best,
Guto

   
AIMLESS Summary
   Overall  InnerShell  OuterShell
Low resolution limit   27.75 27.75  1.07
High resolution limit   1.05  5.75  1.05

Rmerge  (within I+/I-) 0.050 0.078 0.466
Rmerge  (all I+ and I-)0.051 0.080 0.536
Rmeas (within I+/I-)   0.055 0.086 0.591
Rmeas (all I+ & I-)0.054 0.085 0.613
Rpim (within I+/I-)0.023 0.034 0.359
Rpim (all I+ & I-) 0.017 0.028 0.288
Rmerge in top intensity bin0.049- - 
Total number of observations  107950   779  1972
Total number unique1131588   486
Mean((I)/sd(I)) 19.7  46.2   1.8
Mn(I) half-set correlation CC(1/2) 0.998 0.994 0.796
Completeness99.1  99.2  90.4
Multiplicity 9.5   8.9   4.1

Anomalous completeness  98.1 100.0  79.1
Anomalous multiplicity   5.0   6.4   2.2
DelAnom correlation between half-sets -0.067 0.286 0.097
Mid-Slope of Anom Normal Probability   0.789   - -  

Estimate of maximum resolution for significant anomalous signal =  1.14A, from 
CCanom >  0.15

Estimates of resolution limits: overall
   from half-dataset correlation CC(1/2) >  0.30: limit =  1.05A  == maximum 
resolution
   from Mn(I/sd) >  1.50: limit =  1.05A  == maximum 
resolution
   from Mn(I/sd) >  2.00: limit =  1.07A 

Estimates of resolution limits in reciprocal lattice directions:
  Along h axis
   from half-dataset correlation CC(1/2) >  0.30: limit =  1.06A 
   from Mn(I/sd) >  1.50: limit =  1.09A 
  Along k axis
   from half-dataset correlation CC(1/2) >  0.30: limit =  1.11A 
   from Mn(I/sd) >  1.50: limit =  1.13A 
  Along l axis
   from half-dataset correlation CC(1/2) >  0.30: limit =  1.05A  == maximum 
resolution
   from Mn(I/sd) >  1.50: limit =  1.05A 

Anisotropic deltaB (i.e. range of principal components), A^2:  6.40

Average unit cell:   29.12   29.26   55.50   90.00   90.00   90.00 
Space group: I 2 2 2
Average mosaicity:   0.36

AIMLESS Laue Group prediction

   Laue GroupLklhd   NetZc  Zc+   Zc-CCCC-  Rmeas   R-  Delta 
ReindexOperator

= 1I m m m  ***  0.987   6.25  9.19  2.94   0.92  0.29   0.07  0.49   0.0 
[h,k,l]
  2  I 1 2/m 1   0.004   4.36  9.32  4.96   0.93  0.50   0.07  0.30   0.0 
[-h,-k,l]
  3  I 1 2/m 1   0.004   4.14  9.24  5.10   0.92  0.51   0.07  0.30   0.0 
[k,-h,l]
  4  I 1 2/m 1   0.004   4.05  9.23  5.18   0.92  0.52   0.06  0.31   0.0 
[h,-l,k]
  5  I 4/m m m   0.000   6.35  6.35  0.00   0.64  0.00   0.22  0.00   0.3 
[h,k,l]
  6  I 4/m   0.000   0.90  6.83  5.93   0.68  0.59   0.17  0.26   0.3 
[h,k,l]
  7   P -1   0.000   3.69  9.36  5.67   0.94  0.57   0.06  0.26   0.0 
[-h,k,1/2h-1/2k-1/2l]
  8  I 1 2/m 1   0.000   0.08  6.40  6.33   0.64  0.63   0.23  0.22   0.3 
[-1/2h+1/2k-1/2l,-h-k,-1/2h+1/2k+1/2l]
  9F m m m   0.000  -0.43  6.17  6.60   0.62  0.66   0.24  0.20   0.3 
[h+k,-h+k,l]
 10  I 1 2/m 1   0.000   0.75  6.89  6.14   0.69  0.61   0.22  0.22   0.3 
[-1/2h-1/2k-1/2l,h-k,-1/2h-1/2k+1/2l]


Xtriage Summary

Twinning and intensity statistics summary (acentric data):

Statistics independent of twin laws
  /^2 : 2.126  (untwinned: 2.0, perfect twin: 1.5)
  ^2/ : 0.774  (untwinned: 0.785, perfect twin: 0.885)
  <|E^2-1|>   : 0.761  (untwinned: 0.736, perfect twin: 0.541)
  <|L|>, : 0.490, 0.323
  Multivariate Z score L-test: 1.303

 The multivariate Z score is a quality measure of the given
 

[ccp4bb] Job Opening at Albert Einstein College of Medicine

[Syun-Ru Yeh]

A Postdoctoral Position in Structural Biology, Enzymology and Drug Development

A position for a NIH-funded postdoctoral fellow is available immediately in the 
Syun-Ru Yeh Laboratory at Albert Einstein College of Medicine. The postdoctoral 
fellow will take a lead in (1) elucidating structure and function relationships 
of two immunotherapeutic drug targets, human indoleamine 2,3-dioxygenase and 
tryptophan dioxygenase, and (2) developing inhibitors targeting the enzymes as 
therapeutic strategies for cancer treatment. The postdoctoral fellow will join 
an exciting program with a focus on enzymology, spectroscopy-guided 
crystallography and inhibitor development.

The successful applicant must have demonstrated experience in chemistry, 
biochemistry, biophysics or closely related field. Expertise in x-ray 
crystallography, spectroscopy and/or computational modeling is a plus. 
Applicants should provide a CV and a cover letter justifying their suitability 
and interest in the position. Applicants should also arrange to have three 
letters of reference sent to Dr. Syun-Ru Yeh (syun-ru@einstein.yu.edu).

Albert Einstein College of Medicine is located in New York in a scenic Bronx 
campus that includes comfortable, affordable housing in an area rich with 
recreation and entertainment. The scientific environment provides outstanding 
research training opportunities.





To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

[ccp4bb] AW: [EXTERNAL] [ccp4bb] Weird diffraction pattern

[Herman . Schreuder]

To me, it looks like some intergrown salt crystal.
HS

Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Sam Tang
Gesendet: Dienstag, 9. Oktober 2018 13:13
An: CCP4BB@JISCMAIL.AC.UK
Betreff: [EXTERNAL] [ccp4bb] Weird diffraction pattern

Dear all

Hello. We recently shot a crystal (a protein with small molecule as ligand) at 
a synchrotron source and see a weird pattern. 
(https://drive.google.com/file/d/11bEtTJzKaAB5ZybezgN1cqBrckSRg2OV/view?usp=sharing)

Crystal was grown in Citric acid and ammonium sulfate, cryoprotected with 
glycerol.

At first we thought it was a protein crystal contaminated with salt but on 
second thought, the lowest resolution spot was at around 7 A, which doesn't 
make sense for a protein. So we would like to solicit your experience and 
perhaps someone may have encountered similar pattern before?

Many thanks.

Kind regards

Sam




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

Re: [ccp4bb] Weird diffraction pattern

[colin.n...@diamond.ac.uk]

Hello again
It could be a powder pattern aligned along one axis?
The cell I gave (actually orthorhombic) is one of the crystal forms of one of 
the components in your mixture – glycerol. You may have another form of 
glycerol. It is worth checking.

Colin
PS I think glycerol diffraction has been raised previously on CCP4bb


From: Sam Tang 
Sent: 09 October 2018 15:02
To: Nave, Colin (DLSLtd,RAL,LSCI) 
Cc: ccp4bb 
Subject: Re: [ccp4bb] Weird diffraction pattern

Hello Colin

Although the unit cell dimensions from mosflm should be largely unreliable in 
this case, the software actually returned a P2 space group with a=24.6, b=7.5, 
c=69.5  where b is so short that it resembles a small molecule crystal.

Regards

Sam
Sam

On Tue, 9 Oct 2018 at 20:53, 
colin.n...@diamond.ac.uk 
mailto:colin.n...@diamond.ac.uk>> wrote:
Sam
Would this unit cell index some of the spots?
a = 7.00 ± 0.04 A, b = 9.96 ± 0.05 A, c = 6.29 ± 0.04 A.
  Colin

From: CCP4 bulletin board mailto:CCP4BB@JISCMAIL.AC.UK>> 
On Behalf Of Sam Tang
Sent: 09 October 2018 12:13
To: ccp4bb mailto:ccp4bb@jiscmail.ac.uk>>
Subject: [ccp4bb] Weird diffraction pattern

Dear all

Hello. We recently shot a crystal (a protein with small molecule as ligand) at 
a synchrotron source and see a weird pattern. 
(https://drive.google.com/file/d/11bEtTJzKaAB5ZybezgN1cqBrckSRg2OV/view?usp=sharing)

Crystal was grown in Citric acid and ammonium sulfate, cryoprotected with 
glycerol.

At first we thought it was a protein crystal contaminated with salt but on 
second thought, the lowest resolution spot was at around 7 A, which doesn't 
make sense for a protein. So we would like to solicit your experience and 
perhaps someone may have encountered similar pattern before?

Many thanks.

Kind regards

Sam




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1



--

This e-mail and any attachments may contain confidential, copyright and or 
privileged material, and are for the use of the intended addressee only. If you 
are not the intended addressee or an authorised recipient of the addressee 
please notify us of receipt by returning the e-mail and do not use, copy, 
retain, distribute or disclose the information in or attached to the e-mail.
Any opinions expressed within this e-mail are those of the individual and not 
necessarily of Diamond Light Source Ltd.
Diamond Light Source Ltd. cannot guarantee that this e-mail or any attachments 
are free from viruses and we cannot accept liability for any damage which you 
may sustain as a result of software viruses which may be transmitted in or with 
the message.
Diamond Light Source Limited (company no. 4375679). Registered in England and 
Wales with its registered office at Diamond House, Harwell Science and 
Innovation Campus, Didcot, Oxfordshire, OX11 0DE, United Kingdom




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

Re: [ccp4bb] Weird diffraction pattern

[Sam Tang]

Hello Colin

Although the unit cell dimensions from mosflm should be largely unreliable
in this case, the software actually returned a P2 space group with a=24.6,
b=7.5, c=69.5  where b is so short that it resembles a small molecule
crystal.

Regards

Sam
Sam


On Tue, 9 Oct 2018 at 20:53, colin.n...@diamond.ac.uk <
colin.n...@diamond.ac.uk> wrote:

> Sam
>
> Would this unit cell index some of the spots?
>
> a = 7.00 ± 0.04 A, b = 9.96 ± 0.05 A, c = 6.29 ± 0.04 A.
>
>   Colin
>
>
>
> *From:* CCP4 bulletin board  *On Behalf Of *Sam
> Tang
> *Sent:* 09 October 2018 12:13
> *To:* ccp4bb 
> *Subject:* [ccp4bb] Weird diffraction pattern
>
>
>
> Dear all
>
>
>
> Hello. We recently shot a crystal (a protein with small molecule as
> ligand) at a synchrotron source and see a weird pattern. (
> https://drive.google.com/file/d/11bEtTJzKaAB5ZybezgN1cqBrckSRg2OV/view?usp=sharing
> )
>
>
>
> Crystal was grown in Citric acid and ammonium sulfate, cryoprotected with
> glycerol.
>
>
>
> At first we thought it was a protein crystal contaminated with salt but on
> second thought, the lowest resolution spot was at around 7 A, which doesn't
> make sense for a protein. So we would like to solicit your experience and
> perhaps someone may have encountered similar pattern before?
>
>
>
> Many thanks.
>
>
>
> Kind regards
>
>
> Sam
>
>
>
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
>
>
>
> --
>
> This e-mail and any attachments may contain confidential, copyright and or
> privileged material, and are for the use of the intended addressee only. If
> you are not the intended addressee or an authorised recipient of the
> addressee please notify us of receipt by returning the e-mail and do not
> use, copy, retain, distribute or disclose the information in or attached to
> the e-mail.
> Any opinions expressed within this e-mail are those of the individual and
> not necessarily of Diamond Light Source Ltd.
> Diamond Light Source Ltd. cannot guarantee that this e-mail or any
> attachments are free from viruses and we cannot accept liability for any
> damage which you may sustain as a result of software viruses which may be
> transmitted in or with the message.
> Diamond Light Source Limited (company no. 4375679). Registered in England
> and Wales with its registered office at Diamond House, Harwell Science and
> Innovation Campus, Didcot, Oxfordshire, OX11 0DE, United Kingdom
>
>



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

Re: [ccp4bb] Weird diffraction pattern

[colin.n...@diamond.ac.uk]

Sam
Would this unit cell index some of the spots?
a = 7.00 ± 0.04 A, b = 9.96 ± 0.05 A, c = 6.29 ± 0.04 A.
  Colin

From: CCP4 bulletin board  On Behalf Of Sam Tang
Sent: 09 October 2018 12:13
To: ccp4bb 
Subject: [ccp4bb] Weird diffraction pattern

Dear all

Hello. We recently shot a crystal (a protein with small molecule as ligand) at 
a synchrotron source and see a weird pattern. 
(https://drive.google.com/file/d/11bEtTJzKaAB5ZybezgN1cqBrckSRg2OV/view?usp=sharing)

Crystal was grown in Citric acid and ammonium sulfate, cryoprotected with 
glycerol.

At first we thought it was a protein crystal contaminated with salt but on 
second thought, the lowest resolution spot was at around 7 A, which doesn't 
make sense for a protein. So we would like to solicit your experience and 
perhaps someone may have encountered similar pattern before?

Many thanks.

Kind regards

Sam




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

-- 
This e-mail and any attachments may contain confidential, copyright and or 
privileged material, and are for the use of the intended addressee only. If you 
are not the intended addressee or an authorised recipient of the addressee 
please notify us of receipt by returning the e-mail and do not use, copy, 
retain, distribute or disclose the information in or attached to the e-mail.
Any opinions expressed within this e-mail are those of the individual and not 
necessarily of Diamond Light Source Ltd. 
Diamond Light Source Ltd. cannot guarantee that this e-mail or any attachments 
are free from viruses and we cannot accept liability for any damage which you 
may sustain as a result of software viruses which may be transmitted in or with 
the message.
Diamond Light Source Limited (company no. 4375679). Registered in England and 
Wales with its registered office at Diamond House, Harwell Science and 
Innovation Campus, Didcot, Oxfordshire, OX11 0DE, United Kingdom




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

[ccp4bb] Job Opening at Diamond Light Source

[david.h...@diamond.ac.uk]

Dear All

We are recruiting a senior support scientist to join the MX group at Diamond 
Light Source. The role involves developing and supporting the very active MX 
groups user programme, user support for our international user community, 
training and providing operational support to the group. Opportunities exist to 
work with us on developing automated systems for data collection and associated 
supporting infrastructure.

For further information please contact myself directly. For more details on the 
role and to apply please see:

https://vacancies.diamond.ac.uk/vacancy/mx-village-senior-support-scientist-363207.html

Closing date for applications is Sunday - 14th October.

Best regards

Dave Hall
MX Science Group Leader
Diamond Light Source



-- 
This e-mail and any attachments may contain confidential, copyright and or 
privileged material, and are for the use of the intended addressee only. If you 
are not the intended addressee or an authorised recipient of the addressee 
please notify us of receipt by returning the e-mail and do not use, copy, 
retain, distribute or disclose the information in or attached to the e-mail.
Any opinions expressed within this e-mail are those of the individual and not 
necessarily of Diamond Light Source Ltd. 
Diamond Light Source Ltd. cannot guarantee that this e-mail or any attachments 
are free from viruses and we cannot accept liability for any damage which you 
may sustain as a result of software viruses which may be transmitted in or with 
the message.
Diamond Light Source Limited (company no. 4375679). Registered in England and 
Wales with its registered office at Diamond House, Harwell Science and 
Innovation Campus, Didcot, Oxfordshire, OX11 0DE, United Kingdom




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1