Re: [ccp4bb] A quick question about making PEG cryo-protectant

[Jan]

Hi Xiao,
what you run into here is a aqueous two phase system, phosphate and PEG tend to 
form that, see wikipedia entry.

35% PEG 3350 is not a bad cryo, the combination with phosphate is just 
unfortunate. Any chance you can replace with a different salt or buffer? I 
don't know if adding small PEGs will help.
Glycerol, MPD, sugars or oils might be helpful.

Cheers,
Jan
--
Jan Abendroth
Beryllium
Seattle / Bainbridge Island WA, USA
home: Jan.Abendroth_at_gmail.com
work: JAbendroth_at_be4.com
http://www.be4.com

On Oct 13, 2014, at 6:08 PM, Xiao Xiao  wrote:

> Hi Mayer and Jason,
> 
> Thanks for your quick reply! I read from Hampton that for PEG smaller than 
> 5K, directly increasing its concentration is a way of making cryo. But you're 
> right, glycerol or PEG400 will be a good way and might be easier.  I'll try 
> that first.
> 
> Thanks a lot!
> 
> Best,
> Xiao
> 
> 2014-10-13 17:59 GMT-07:00 Mayer, Mark (NIH/NICHD) [E] :
> Try either glycerol or PEG 400.
> Increasing concentration of 'high' MW PEG is not a good strategy.
> 
> 
> From: Xiao Xiao [victor41...@gmail.com]
> Sent: Monday, October 13, 2014 8:51 PM
> To: CCP4BB@JISCMAIL.AC.UK
> Subject: [ccp4bb] A quick question about making PEG cryo-protectant
> 
> Hi,
> 
> Since my crystallization condition contains PEG, I am trying to use PEG as 
> one of my cryo for testing. For example, one crystallization condition is 
> 0.2M Na2PO4, 20%PEG3350. I want to make 0.2M Na2PO4 and 40% PEG3350 as 
> cryoprotectant. When I added 200ul salt from 1M stock, and 800ul PEG from 50% 
> stock, mixed them, seems they stayed as emulsion (confirmed by microscope), 
> no matter how I pipette or vortex them. I tried to incubate it at 40degree, 
> seems it got worse.
> 
> I am now trying to re-make the cryo from solid reagent, not solution stock. 
> Will this be helpful? Or is there any other trick?
> 
> I will really appreciate!
> 
> Best,
> Xiao
> 

[ccp4bb] Post Doctoral Position - University of Oklahoma

[Thomas, Leonard M.]

The following position is available at the University of Oklahoma.  Please 
contact Liz Karr for any addition information.

A Postdoc position is available in microbiology and structural biology in the
research group of Dr. Liz Karr in the Department of Microbiology and Plant 
Biology at the University of Oklahoma, Norman OK. The lab is part of an NIH 
COBRE in Structural Biology as well as a member of the Michael F. Price 
Institute for Structural Biology (ISB) housed in the Department of Chemistry 
and Biochemistry where there is an emphasis in anaerobic structural biology.
Our group studies structure-function relationships of microbial transcription 
regulators. The postdoc will focus on studies of the novel methanogen specific 
transcription regulator, MsvR first identified by the Karr Lab. MsvR is a 
redox- sensitive transcription regulator that employs a number of cysteine 
residues to sense the redox environment in strict anaerobes. The project 
involves crystallization of MsvR and the MsvR/DNA complex and characterization 
of MsvR variants in vivo and in vitro. The lab utilizes a genetically tractable 
methanogen to study the in vivo role of MsvR in cell survival and to assess 
changes in the host transcriptome in msvR deletion strains. On a regular basis 
the postdoc will be involved in cloning, protein overexpression, site directed 
mutagenesis, anaerobic culturing and genetic manipulations as well as aerobic 
and anaerobic protein crystallization.
We are looking for creative and motivated applicants with a strong background 
in molecular genetics and anaerobic microbiology and/or structural biology. The 
candidate must provide evidence of their research potential through 
publications and presentations, and activities involving mentoring and lab 
management are an asset.
The initial appointment is for one year and is renewable upon satisfactory 
progress and continued funding. Salary level commensurate with experience. 
Equal opportunity, applications from under-represented groups are encouraged.
Applications and informal inquiries should be sent to Liz Karr by email at 
lizkarr (at) ou (dot) edu. Applications should contain a letter describing your 
motivation and career goals, a CV highlighting your important contributions to 
research activities, and the contact details for three professional references. 
Applications will be accepted until the position is filled.



Leonard M. Thomas Ph.D.
Macromolecular Crystallography Laboratory Manager
University of Oklahoma
Department of Chemistry and Biochemistry
Stephenson Life Sciences Research Center
101 Stephenson Parkway
Norman, OK 73019
405-325-1126
lmtho...@ou.edu
http://barlywine.chem.ou.edu
http://structuralbiology.ou.edu

Re: [ccp4bb] A quick question about making PEG cryo-protectant

[Xiao Xiao]

Hi Mayer and Jason,

Thanks for your quick reply! I read from Hampton that for PEG smaller than
5K, directly increasing its concentration is a way of making cryo. But
you're right, glycerol or PEG400 will be a good way and might be easier.
I'll try that first.

Thanks a lot!

Best,
Xiao

2014-10-13 17:59 GMT-07:00 Mayer, Mark (NIH/NICHD) [E] 
:

> Try either glycerol or PEG 400.
> Increasing concentration of 'high' MW PEG is not a good strategy.
>
> 
> From: Xiao Xiao [victor41...@gmail.com]
> Sent: Monday, October 13, 2014 8:51 PM
> To: CCP4BB@JISCMAIL.AC.UK
> Subject: [ccp4bb] A quick question about making PEG cryo-protectant
>
> Hi,
>
> Since my crystallization condition contains PEG, I am trying to use PEG as
> one of my cryo for testing. For example, one crystallization condition is
> 0.2M Na2PO4, 20%PEG3350. I want to make 0.2M Na2PO4 and 40% PEG3350 as
> cryoprotectant. When I added 200ul salt from 1M stock, and 800ul PEG from
> 50% stock, mixed them, seems they stayed as emulsion (confirmed by
> microscope), no matter how I pipette or vortex them. I tried to incubate it
> at 40degree, seems it got worse.
>
> I am now trying to re-make the cryo from solid reagent, not solution
> stock. Will this be helpful? Or is there any other trick?
>
> I will really appreciate!
>
> Best,
> Xiao
>

[ccp4bb] A quick question about making PEG cryo-protectant

[Xiao Xiao]

Hi,

Since my crystallization condition contains PEG, I am trying to use PEG as
one of my cryo for testing. For example, one crystallization condition is
0.2M Na2PO4, 20%PEG3350. I want to make 0.2M Na2PO4 and 40% PEG3350 as
cryoprotectant. When I added 200ul salt from 1M stock, and 800ul PEG from
50% stock, mixed them, seems they stayed as emulsion (confirmed by
microscope), no matter how I pipette or vortex them. I tried to incubate it
at 40degree, seems it got worse.

I am now trying to re-make the cryo from solid reagent, not solution stock.
Will this be helpful? Or is there any other trick?

I will really appreciate!

Best,
Xiao

[ccp4bb] incorrect R-factor calculation in sftools

[Tim Gruene]

Dear all (dear developers),

on a recent discussion on the phenixbb, Nat figured out that sftools
calculates the R-factor incorrectly as

"200*Sum|col1-col2|/sum(col1+col2)"

instead of

"100*Sum|col1-col2|/sum(col1)"

May I suggest to either correct this or not call it Rfactor in order to
avoid future confusion (as it did on the phenibb)?

Best wishes,
Tim
-- 
Dr Tim Gruene
Institut fuer anorganische Chemie
Tammannstr. 4
D-37077 Goettingen

GPG Key ID = A46BEE1A



signature.asc
Description: OpenPGP digital signature

Re: [ccp4bb] experimental phasing + MR

[Gabriel Salzman]

I appreciate all of your responses and I will give your recommendations a try!


Thanks,
Gabriel


On Oct 13, 2014, at 1:50 PM, Kay Diederichs  
wrote:

> Hi Gabriel,
> 
> what you want to do is called MRSAD - use the partial MR model to find the 
> sites, combine the phase information, and complete the model. There are hits 
> with the keyword MRSAD if you search on phenix-online.org
> 
> HTH,
> Kay

Re: [ccp4bb] experimental phasing + MR

[Jason Busby]

I’ve had success with MRSAD as Kay suggests.  One option is to try putting 
everything into Auto-Rickshaw.  It can run multiple cycles of MRSAD, 
auto-building a model for improved molecular replacement, finding new heavy 
atom sites and repeating.  

There is a paper on this available:  http://dx.doi.org/10.1107/S0907444909029643
And an Auto-Rickshaw server:  http://www.embl-hamburg.de/Auto-Rickshaw/

Cheers,

Jason.

—
Dr Jason Busby
Laboratory of Structural Biology
School of Biological Sciences
The University of Auckland
Private Bag 92019
Auckland Mail Centre
Auckland
New Zealand

ph:  +64 9 3737599 ext 88958

On 14/10/2014, at 6:48 am, Gabriel Salzman  wrote:

> Hello,
> 
> I am wondering if anyone has experience or advice for my current situation:
> 
> I have a native dataset of a ~50kD protein complex at 2.4angstroms. One of 
> the members of the complex (~10kD) was able to be found easily by molecular 
> replacement. The space group was determined as P65. I was able to use 
> phenix.mr_rosetta to generate a partial model of the other member of the 
> complex (~half of this 40kD protein). When I do molecular replacement for 
> these two members of the complex in PHASER, the TFZ score is ~20-30. 
> Therefore, I am confident that this partial model is accurate.
> 
> I recently soaked some complex crystals in a variety of heavy atoms to obtain 
> experimental phase information. My best signal is from an iodide soak. These 
> crystals diffracted to 3 angstroms and I can see a very weak but significant 
> anomalous signal to 5-4.5 angstroms. I have used autoSHARP to determine that 
> I have one site of ~99% occupancy and 4 other sites of <50% occupancy. I am 
> inclined to think these other sites are not true. My density modified map 
> seems to me to be better if I only include the single high-occupancy site in 
> the phase determination. Even with this weak experimental phase information, 
> my map is not good enough to see the substantial density from the piece of my 
> protein that was not included in the MR model. 
> 
> I have also tried phenix.phaser_EP with mediocre results.
> 
> I apologize for the long question, and I would greatly appreciate any 
> suggestions for how to combine all of these data into a workable map so I can 
> build out the rest of the structure.
> 
> Thanks,
> Gabriel

[ccp4bb] Postdoctoral position -Structural and Chemical biology

[Jinrong Min]

A Post-doctoral position in structural and chemical biology is available in the 
Chromatin structural biology and Epigenetic group
at the Structural Genomics Consortium, University of Toronto. My lab aims to 
characterize chromatin modifying enzymes and chromatin modification recognition 
proteins by X-ray crystallography in combination with other biochemical and 
biophysical techniques. Please check our lab website for more details:  
http://www.sgc.utoronto.ca/min.


Applicants should hold a Ph.D. in  molecular biology, biochemistry  or 
structural biology with 0-3 years  postdoctoral
research experience; prior training in X-ray crystallography is desirable, but 
not necessary. Interested candidates are
invited to send their CV with 3 references to jr@utoronto.ca.

Jinrong Min, PhD
Principal Investigator, Chromatin Structural Biology
Structural Genomics Consortium
Associate Professor, Department of Physiology
University of Toronto
101 College St., Toronto
On M5G 1L7, Canada
Tel: 416-9463868
Email:  jr@utoronto.ca

[ccp4bb] Industrial Position of Scientist Structural Biology at Amgen

[Xin Huang]

Dear All,

We are looking for a scientist to join us in the area of structural biology of 
membrane proteins.
Please see the attached file for a description of the job and apply at 
http://careers.amgen.com/en/jobs/descriptions/scientist-structural-biology-cambridge-massachusetts-job-4826930.

Best regards,
Xin


Sci Structural Biology AMA.pdf
Description: Adobe PDF document

Re: [ccp4bb] experimental phasing + MR

[Kay Diederichs]

Hi Gabriel,

what you want to do is called MRSAD - use the partial MR model to find the 
sites, combine the phase information, and complete the model. There are hits 
with the keyword MRSAD if you search on phenix-online.org

HTH,
Kay

[ccp4bb] experimental phasing + MR

[Gabriel Salzman]

Hello,

I am wondering if anyone has experience or advice for my current situation:

I have a native dataset of a ~50kD protein complex at 2.4angstroms. One of the 
members of the complex (~10kD) was able to be found easily by molecular 
replacement. The space group was determined as P65. I was able to use 
phenix.mr_rosetta to generate a partial model of the other member of the 
complex (~half of this 40kD protein). When I do molecular replacement for these 
two members of the complex in PHASER, the TFZ score is ~20-30. Therefore, I am 
confident that this partial model is accurate.

I recently soaked some complex crystals in a variety of heavy atoms to obtain 
experimental phase information. My best signal is from an iodide soak. These 
crystals diffracted to 3 angstroms and I can see a very weak but significant 
anomalous signal to 5-4.5 angstroms. I have used autoSHARP to determine that I 
have one site of ~99% occupancy and 4 other sites of <50% occupancy. I am 
inclined to think these other sites are not true. My density modified map seems 
to me to be better if I only include the single high-occupancy site in the 
phase determination. Even with this weak experimental phase information, my map 
is not good enough to see the substantial density from the piece of my protein 
that was not included in the MR model. 

I have also tried phenix.phaser_EP with mediocre results.

I apologize for the long question, and I would greatly appreciate any 
suggestions for how to combine all of these data into a workable map so I can 
build out the rest of the structure.

Thanks,
Gabriel

Re: [ccp4bb] Refinement of Iron-sulphur clusters

[Pedro Matias]

Deat CCP4ers,

In relation to this topic, I'd like to mention that SF4 has replaced FS4
as the Fe4S4 monomer in the CCP4 monomer library.

However, the dictionary values for bond lengths and angles are not
correct, and this is especially noticeable when the dictionary is used
in a high-resolution refinement (e.g., 1.05 A). In particular, the
Fe-S-Fe angles are all set to 90 degrees, when in fact some are smaller
and others are larger.

This introduces a bias in the final rms angle deviation values. Fe4S4 is
a distorted cubane, not a perfect cube.

Best regards,

Pedro Matias

Em 13-10-2014 12:09, "Oliver Smart" escreveu:
> Stephen,
>
> Robbie advice is 100% correct. Be careful about the naming of the sulphur
> atoms (is S1 opposite S4 or next to it). I recall that the distributed CCP4
> dictionaries have different atom naming than the PDB chemical components
> dictionary.  If this is the problem is that the naming is different this
> should result
> in large restraint violations that should be appear in the REFMAC output
> (others can tell you where).
>
> In general diffraction from Fe S clusters is so strong that the atom
> positions
> are pretty much determined from the electron density. But in case it helps
> I did some work data mining restraints for Fe2S2 and Fe4S4 from CSD
> structures (this has to be done manually). Please find attached the
> resulting FES.cif and SF4.cif restraint dictionaries (distributed with
> BUSTER).
>
> Good luck,
>
> Oliver
> ---
> Dr Oliver Smart
> Director SmartSci Limited http://www.smartsci.uk/
> & Consultant Global Phasing Ltd http://www.globalphasing.com/
>
> on 10/10/14 5:33 PM, Robbie Joosten  wrote:
>
>> Dear Stephen,
>>
>> The dictionary is very specific about atom names. If you have them swapped
>> (as many PDB entries do). The angle and chiral volume restraints will
> wreck
>> your cluster. You also need to make sure to provide LINK records to attach
>> the cluster to the surrounding cysteines.
>>
>> HTH,
>> Robbie
>>
>> Sent from my Windows Phone
>> 
>> Van: 
>> Verzonden: 10-10-2014 18:07
>> Aan: CCP4BB@JISCMAIL.AC.UK
>> Onderwerp: [ccp4bb] Refinement of Iron-sulphur clusters
>>
>> Dear CCP4 BBers,
>>
>> I am currently refining a model of a protein containing three iron sulphur
>> clusters using Refmac (v5.8.0078) and am getting some unusual results.
> One
>> of the clusters (3fe 4s) seems to behave resonably and the refined
> electron
>> density looks very nice.  The atoms in the others clusters (4Fe4S and
>> 4Fe3S), however, migrate towards the centre of the cluster producing Fo-Fc
>> difference maps with strong negative density at the centre surrounded by
>> blobs of positive density where the atoms should be.
>>
>> I have checkedthe refmac dictionaries and there are entries for each of
> the
>> clusters, so I shouldn't have to explicitly include a cif file for the
>> clusters as an input right?  In fact, when I did try to include a
> dictionary
>> file for one of the clusters refmac gave a warning message in the log
> about
>> duplicate monomers dictionaries and it made no difference to the
>> refinement/maps anyway.
>>
>> On checking the refmac log file there is no specific mention that there
> are
>> hetero-groups in the pdb file and no explicit mention that it is using any
>> dictionaries during the refinement.  How can I tell if refmac is
>> reading/using the dictionaries?  Also bonds within the clusters are
> flagged
>> up as outliers and deviating by more than 10 sigma from ideal suggesting
>> that they are not getting read?
>>
>> I realise that something chemically odd could be going on at the centres,
>> but if I omit them from the model and calculate maps very strong density
>> comes back around where the atoms sat before refinement, so I am pretty
>> confident that they are in the right place to start with.
>>
>> Any help with this problem would be greatly appreciated, I'm sure there is
>> something obvious that I am missing but can't see what that is at the
>> moment.  It is particularly confusing since one cluster apparently behaves
>> while the others do not.  I could, of course, try phenix or buster, but I
>> would like to get to the bottom of the problem with refmac if possible.
>>
>> Thanks very much in advance,
>>
>> Steve
>>
>> Dr Stephen Carr
>> Research Complex at Harwell (RCaH)
>> Rutherford Appleton Laboratory
>> Harwell Oxford
>> Didcot
>> Oxon OX11 0FA
>> United Kingdom
>> Email stephen.c...@rc-harwell.ac.uk
>> tel 01235 567717
>> This email 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 authorized 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
>> this email.
>>
>> Any views or opinions presented are solely those of the au

[ccp4bb] Free seminar on Characterising Biomolecular Interactions - 23rd October

[Mahey, Jas]

TA Instruments invites you to:
Characterising Biomolecular Interactions
Complementary Seminar Presented
by TA Instruments



This seminar includes a range of topics and will cover techniques available for 
the characterisation of Biomolecular interactions - with a focus on the use of 
Calorimetric tools such as ITC and DSC.

Attendance is free, but as numbers will be limited registration will be offered 
on a first come first serve basis.

LUNCH will be provided and the meeting will conclude with a free tour of the 
Williams F1 collection.
Agenda

Studying Biomolecular Interactions by ITC
- John Ladbury, MD Anderson Cancer Centre, University of Texas
Protein-protein interactions in membrane trafficking
- David Owen, Cambridge Institute for Medical Research, Cambridge University
ITC for Enzyme Kinetics
- Malin Suurkuusk, TA Instruments
DSC Characterization of the Structure/Function Relationship for Proteins
- Dile Holton, TA Instruments
Studying interactions between protein, small molecules and water by DSC
- Robert Falconer, Chemical & Biological Engineering, Sheffield University
ITC and DSC data analysis -Tips and Tricks on Model Selection
- Malin Suurkuusk, TA Instruments
Free Tour of Williams F1 Museum


 Complete Info


 Register Now


When
October 23rd 2014
09:00 - 16:30

What
Characterising Biomolecular Interactions

Where
The Williams Conference Centre, Wantage
(near Oxford)

Fee
There is no charge for this event, but advance registration is required to 
reserve your spot. Space is limited so register early.

More Information
Contact Emma Donnachie
TA Instruments
+44 208 238 6126
edonnac...@tainstruments.com

Quick Links
Register Now






watzip



===
The information in this email is confidential, and is intended solely for the 
addressee(s).
Access to this email by anyone else is unauthorized and therefore prohibited. 
If you are
not the intended recipient you are notified that disclosing, copying, 
distributing or taking
any action in reliance on the contents of this information is strictly 
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===

Re: [ccp4bb] Refinement of Iron-sulphur clusters

Dear Matthew, Robbie Hans and Oliver,

Thanks for the advice, it seems that it is simply the geometry definitions in 
the dictionaries distributed with CCP4 6.4 are different to those calculated 
from the atomic coordinates.  For example, the chiral volume definitions in the 
for the 4Fe4S cluster are the opposite sign to those in my protein.  So 
swapping the atom names has sorted that, I am still working on the other 
clusters, but progress is being made.

Since I solved the structure by MR using coordinates downloaded from the pdb, I 
assumed the atom names would agree with the geometry definitions in Refmac 
(since the search model was also refined with refmac).  Have the cif 
dictionaries been modified/updated recently? 

thanks again,

Steve 

Dr Stephen Carr
Research Complex at Harwell (RCaH)
Rutherford Appleton Laboratory
Harwell Oxford
Didcot
Oxon OX11 0FA
United Kingdom
Email stephen.c...@rc-harwell.ac.uk
tel 01235 567717


From: "Oliver Smart" [osm...@smartsci.uk]
Sent: 13 October 2014 12:09
To: ccp4bb
Subject: Re: [ccp4bb] Refinement of Iron-sulphur clusters

Stephen,

Robbie advice is 100% correct. Be careful about the naming of the sulphur
atoms (is S1 opposite S4 or next to it). I recall that the distributed CCP4
dictionaries have different atom naming than the PDB chemical components
dictionary.  If this is the problem is that the naming is different this
should result
in large restraint violations that should be appear in the REFMAC output
(others can tell you where).

In general diffraction from Fe S clusters is so strong that the atom
positions
are pretty much determined from the electron density. But in case it helps
I did some work data mining restraints for Fe2S2 and Fe4S4 from CSD
structures (this has to be done manually). Please find attached the
resulting FES.cif and SF4.cif restraint dictionaries (distributed with
BUSTER).

Good luck,

Oliver
---
Dr Oliver Smart
Director SmartSci Limited http://www.smartsci.uk/
& Consultant Global Phasing Ltd http://www.globalphasing.com/

on 10/10/14 5:33 PM, Robbie Joosten  wrote:

> Dear Stephen,
>
> The dictionary is very specific about atom names. If you have them swapped
> (as many PDB entries do). The angle and chiral volume restraints will
wreck
> your cluster. You also need to make sure to provide LINK records to attach
> the cluster to the surrounding cysteines.
>
> HTH,
> Robbie
>
> Sent from my Windows Phone
> 
> Van: 
> Verzonden: 10-10-2014 18:07
> Aan: CCP4BB@JISCMAIL.AC.UK
> Onderwerp: [ccp4bb] Refinement of Iron-sulphur clusters
>
> Dear CCP4 BBers,
>
> I am currently refining a model of a protein containing three iron sulphur
> clusters using Refmac (v5.8.0078) and am getting some unusual results.
One
> of the clusters (3fe 4s) seems to behave resonably and the refined
electron
> density looks very nice.  The atoms in the others clusters (4Fe4S and
> 4Fe3S), however, migrate towards the centre of the cluster producing Fo-Fc
> difference maps with strong negative density at the centre surrounded by
> blobs of positive density where the atoms should be.
>
> I have checkedthe refmac dictionaries and there are entries for each of
the
> clusters, so I shouldn't have to explicitly include a cif file for the
> clusters as an input right?  In fact, when I did try to include a
dictionary
> file for one of the clusters refmac gave a warning message in the log
about
> duplicate monomers dictionaries and it made no difference to the
> refinement/maps anyway.
>
> On checking the refmac log file there is no specific mention that there
are
> hetero-groups in the pdb file and no explicit mention that it is using any
> dictionaries during the refinement.  How can I tell if refmac is
> reading/using the dictionaries?  Also bonds within the clusters are
flagged
> up as outliers and deviating by more than 10 sigma from ideal suggesting
> that they are not getting read?
>
> I realise that something chemically odd could be going on at the centres,
> but if I omit them from the model and calculate maps very strong density
> comes back around where the atoms sat before refinement, so I am pretty
> confident that they are in the right place to start with.
>
> Any help with this problem would be greatly appreciated, I'm sure there is
> something obvious that I am missing but can't see what that is at the
> moment.  It is particularly confusing since one cluster apparently behaves
> while the others do not.  I could, of course, try phenix or buster, but I
> would like to get to the bottom of the problem with refmac if possible.
>
> Thanks very much in advance,
>
> Steve
>
> Dr Stephen Carr
> Research Complex at Harwell (RCaH)
> Rutherford Appleton Laboratory
> Harwell Oxford
> Didcot
> Oxon OX11 0FA
> United Kingdom
> Email stephen.c...@rc-harwell.ac.uk
> tel 01235 567717
> This email and any attachments may contain confidential, copyright and or
> privileged material, 

Re: [ccp4bb] I222 - P22121 space-group ambiguity

[Bernhard Rupp]

Yes, I missed the second set of cell constants and assumed Florian is
talking about ambiguity in the same data set.

 

Thx BR

 

From: Harry Powell [mailto:ha...@mrc-lmb.cam.ac.uk] 
Sent: Montag, 13. Oktober 2014 12:14
To: b...@hofkristallamt.org
Cc: Harry Powell; CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] I222 - P22121 space-group ambiguity

 

Hi Bernhard et al

 



The lattice metric  is the same for both SGs (did not understand Eleanor’s
remark)

 

Hmmm - 

well the I212121 cell and the P21221 cells you give here have different
volumes, so they cant be just rearrangements of symmetry operators and
non-cryst translation.

 

Cell volume for the I222 cell = 87.8 * 101.18 * 123.63 ~= 1,098,280 Å^3

 

Cell volume for the P22121 cell = 93.34 * 105.47 * 122.98 ~=1,210,685 Å^3

 

Or ~10% different - so to me, anyway, Eleanor's remark made perfect sense.





 

In principle, differences between NCS and XS of compatible metric become
more distinct at

higher resolution (i.e. at night, all cats are grey).

 

Best, BR

 

From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of
Florian Schmitzberger
Sent: Montag, 13. Oktober 2014 10:47
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] I222 - P22121 space-group ambiguity

 

Hi everybody,

 

I collected a number of X-ray data sets from crystals originating from the
same cryst. drop. I solved the initial structure in P22121 space group by MR
with Phaser locating two molecules (data to ~ 2.1 Angstr.); refined R/Rfree:
0.213/0.244.

 

Processing of some of the other data sets with XDS/Aimless is consistent
with I222 space group (resolution ~ 2.6 Ang.). I can locate one molecule.
The unit-cell dimensions for I222 and the initial P22121 space groups for
two of the data sets are:

I222: a=87.8 b=101.18 c=123.63; P22121: a=93.34 b=105.47 c=122.98;

 

I superposed the molecule in I222 onto one of the two located for the
initially solved P22121; the orientation of the NCS-related molecule in
P22121 differs from the crystallographic-symmetry related one in I222.
Trying to solve this P22121 data set in I222 with MR, does not result in
high Z scores, and maps do not look good.

 

Some of the data sets that process in I222 to ~ 3 Angstr., I can also solve
in P22121, locating two molecules (differences may not be that clear in this
case, since the resolution is lower).

 

Some other data sets process in P22121 with Aimless; with a substantial
off-origin Patterson peak, indicating translational NCS. For these, Phaser
positions two molecules that are related by crystallographic translational
NCS. These two molecules are crystallographic-symmetry related in the
original P22121 data set. I can also solve these data sets in I222 space
group, with the overall Z score higher than for the P22121 data. 

 

I am uncertain, what the ‘true’ space group for some of my data sets is.
Could it be that for data that process in P22121, but can be solved in I222,
reflections that would indicate I222 space group were not collected?
Alternatively, perhaps what I am seeing is that there is a (gradual)
transition of the crystal lattice (between P22121 and I222 or vice versa),
caused by variation in crystal handling/cooling or exposure to X-rays.

 

It’s relevant to me, because in P22121 space group, a region of the molecule
that is of biological interest makes NCS-related crystal contacts that are
crystallographic-symmetry related in I222.

 

Has anybody observed similar cases? I would appreciate comments.

 

Cheers,

 

Florian

 

 

 

Harry

--

Dr Harry Powell, MRC Laboratory of Molecular Biology, Francis Crick Avenue,
Cambridge Biomedical Campus, Cambridge CB2 0QH

Chairman of International Union of Crystallography Commission on
Crystallographic Computing

Chairman of European Crystallographic Association SIG9 (Crystallographic
Computing) 

 

 

 

 

 

 

 





 

Re: [ccp4bb] I222 - P22121 space-group ambiguity

[Bernhard Rupp]

> As I understand, P22121 should be fine to process (and deposit in the
PDB); and there is no need to reindex to HM convention P21212.

 

No need to, but trivial and then in conformance with the ITC. If we have a
standard listing in ITC, why not use it? Having different symbols/settings 

for the same SG only adds complexity to data mining without providing a
tangible benefit. Ergo, non-parsimonious and fails Occam's razor

which in turn follows form Bayes (Jefferys, W.H. & Berger, J.O. (1991).
American Scientist 80, 64-72. Sharpening Ockham's razor on a Bayesian
strop).

 

Best, BR

 

Numquam ponenda est pluralitas sine necessitate 

Multitude must never be proposed without necessity

 

 

Re: [ccp4bb] Refinement of Iron-sulphur clusters

[Oliver Smart]

Stephen,

Robbie advice is 100% correct. Be careful about the naming of the sulphur
atoms (is S1 opposite S4 or next to it). I recall that the distributed CCP4
dictionaries have different atom naming than the PDB chemical components
dictionary.  If this is the problem is that the naming is different this
should result
in large restraint violations that should be appear in the REFMAC output
(others can tell you where).

In general diffraction from Fe S clusters is so strong that the atom
positions
are pretty much determined from the electron density. But in case it helps
I did some work data mining restraints for Fe2S2 and Fe4S4 from CSD
structures (this has to be done manually). Please find attached the
resulting FES.cif and SF4.cif restraint dictionaries (distributed with
BUSTER).

Good luck,

Oliver
---
Dr Oliver Smart
Director SmartSci Limited http://www.smartsci.uk/
& Consultant Global Phasing Ltd http://www.globalphasing.com/

on 10/10/14 5:33 PM, Robbie Joosten  wrote:

> Dear Stephen,
>
> The dictionary is very specific about atom names. If you have them swapped
> (as many PDB entries do). The angle and chiral volume restraints will
wreck
> your cluster. You also need to make sure to provide LINK records to attach
> the cluster to the surrounding cysteines.
>
> HTH,
> Robbie
>
> Sent from my Windows Phone
> 
> Van: 
> Verzonden: 10-10-2014 18:07
> Aan: CCP4BB@JISCMAIL.AC.UK
> Onderwerp: [ccp4bb] Refinement of Iron-sulphur clusters
>
> Dear CCP4 BBers,
>
> I am currently refining a model of a protein containing three iron sulphur
> clusters using Refmac (v5.8.0078) and am getting some unusual results.
One
> of the clusters (3fe 4s) seems to behave resonably and the refined
electron
> density looks very nice.  The atoms in the others clusters (4Fe4S and
> 4Fe3S), however, migrate towards the centre of the cluster producing Fo-Fc
> difference maps with strong negative density at the centre surrounded by
> blobs of positive density where the atoms should be.
>
> I have checkedthe refmac dictionaries and there are entries for each of
the
> clusters, so I shouldn't have to explicitly include a cif file for the
> clusters as an input right?  In fact, when I did try to include a
dictionary
> file for one of the clusters refmac gave a warning message in the log
about
> duplicate monomers dictionaries and it made no difference to the
> refinement/maps anyway.
>
> On checking the refmac log file there is no specific mention that there
are
> hetero-groups in the pdb file and no explicit mention that it is using any
> dictionaries during the refinement.  How can I tell if refmac is
> reading/using the dictionaries?  Also bonds within the clusters are
flagged
> up as outliers and deviating by more than 10 sigma from ideal suggesting
> that they are not getting read?
>
> I realise that something chemically odd could be going on at the centres,
> but if I omit them from the model and calculate maps very strong density
> comes back around where the atoms sat before refinement, so I am pretty
> confident that they are in the right place to start with.
>
> Any help with this problem would be greatly appreciated, I'm sure there is
> something obvious that I am missing but can't see what that is at the
> moment.  It is particularly confusing since one cluster apparently behaves
> while the others do not.  I could, of course, try phenix or buster, but I
> would like to get to the bottom of the problem with refmac if possible.
>
> Thanks very much in advance,
>
> Steve
>
> Dr Stephen Carr
> Research Complex at Harwell (RCaH)
> Rutherford Appleton Laboratory
> Harwell Oxford
> Didcot
> Oxon OX11 0FA
> United Kingdom
> Email stephen.c...@rc-harwell.ac.uk
> tel 01235 567717
> This email 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 authorized 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
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> this email.
>
> Any views or opinions presented are solely those of the author and do not
> necessarily represent those of the Research Complex at Harwell.
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> There is no guarantee that this email 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
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> We use an electronic filing system. Please send electronic versions of
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>


FES.cif
Description: CIF chemical test


SF4.cif
Description: CIF chemical test

Re: [ccp4bb] I222 - P22121 space-group ambiguity

[Schmitzberger,Florian]

Hi everybody,

Thank you for your comments!

The data sets that index in I222 tend to have shorter a and b axes. I have data 
sets in I222 with a=95.31 b=106 c=125.9 though (still need to look at these in 
detail). These axes are relatively close to those in the original P22121, which 
I processed and refined. Some of the P22121 data sets with strong translational 
NCS, which can be solved in I222 too, have dimensions of a=93.04 b=104.3 
c=123.6; i.e. very similar to the original P22121 that is inconsistent with 
I222.

The off-origin Patterson peak in the P22121 space group with trans. NCS is at 
0.46, 0.50, -0.50 and at 22 % height - relative to the origin peak. This peak 
is also present in the originally solved P22121 (where no substantial 
translational NCS was flagged), with a peak height of 18 %.

Cheers,

Florian

P.S.: As I understand, P22121 should be fine to process (and deposit in the 
PDB); and there is no need to reindex to HM convention P21212.

On Oct 13, 2014, at 11:08 AM, Eleanor Dodson 
mailto:eleanor.dod...@york.ac.uk>> wrote:

Hmmm -
well the I212121 cell and the P21221 cells you give here have different 
volumes, so they cant be just rearrangements of symmetry operators and 
non-cryst translation.

Do you get other sets of cell dimensions for different processing?

Eleanor

On 13 October 2014 09:47, Florian Schmitzberger 
mailto:schmitzber...@crystal.harvard.edu>> 
wrote:
Hi everybody,

I collected a number of X-ray data sets from crystals originating from the same 
cryst. drop. I solved the initial structure in P22121 space group by MR with 
Phaser locating two molecules (data to ~ 2.1 Angstr.); refined R/Rfree: 
0.213/0.244.

Processing of some of the other data sets with XDS/Aimless is consistent with 
I222 space group (resolution ~ 2.6 Ang.). I can locate one molecule. The 
unit-cell dimensions for I222 and the initial P22121 space groups for two of 
the data sets are:
I222: a=87.8 b=101.18 c=123.63; P22121: a=93.34 b=105.47 c=122.98;

I superposed the molecule in I222 onto one of the two located for the initially 
solved P22121; the orientation of the NCS-related molecule in P22121 differs 
from the crystallographic-symmetry related one in I222. Trying to solve this 
P22121 data set in I222 with MR, does not result in high Z scores, and maps do 
not look good.

Some of the data sets that process in I222 to ~ 3 Angstr., I can also solve in 
P22121, locating two molecules (differences may not be that clear in this case, 
since the resolution is lower).

Some other data sets process in P22121 with Aimless; with a substantial 
off-origin Patterson peak, indicating translational NCS. For these, Phaser 
positions two molecules that are related by crystallographic translational NCS. 
These two molecules are crystallographic-symmetry related in the original 
P22121 data set. I can also solve these data sets in I222 space group, with the 
overall Z score higher than for the P22121 data.

I am uncertain, what the ‘true’ space group for some of my data sets is. Could 
it be that for data that process in P22121, but can be solved in I222, 
reflections that would indicate I222 space group were not collected? 
Alternatively, perhaps what I am seeing is that there is a (gradual) transition 
of the crystal lattice (between P22121 and I222 or vice versa), caused by 
variation in crystal handling/cooling or exposure to X-rays.

It’s relevant to me, because in P22121 space group, a region of the molecule 
that is of biological interest makes NCS-related crystal contacts that are 
crystallographic-symmetry related in I222.

Has anybody observed similar cases? I would appreciate comments.

Cheers,

Florian

Re: [ccp4bb] I222 - P22121 space-group ambiguity

[Kay Diederichs]

Hi Florian,

a couple of things come to my mind:
a) you have to be careful in your use of XDS; if you specify space group number 
18 it will use "P 21 21 2" as space group, whereas you seem to use the ordering 
ahttp://strucbio.biologie.uni-konstanz.de/xdswiki/index.php/IDXREF.LP ) to find 
whether there are half-integer indices, and to visually check predictions 
(XDSGUI - see http://strucbio.biologie.uni-konstanz.de/xdswiki/index.php/XDSGUI 
) presents a way to do this easily.
c) According to the ITC vol A, P22121 and I222 have a subgroup-supergroup 
relationship so it is indeed possible that a small change in the packing can 
convert one to the other 
d) ZANUDA is the program to use in case of space-group ambiguity

HTH,

Kay

On Mon, 13 Oct 2014 10:47:18 +0200, Florian Schmitzberger 
 wrote:

>Hi everybody,
>
>I collected a number of X-ray data sets from crystals originating from the 
>same cryst. drop. I solved the initial structure in P22121 space group by MR 
>with Phaser locating two molecules (data to ~ 2.1 Angstr.); refined R/Rfree: 
>0.213/0.244.
>
>Processing of some of the other data sets with XDS/Aimless is consistent with 
>I222 space group (resolution ~ 2.6 Ang.). I can locate one molecule. The 
>unit-cell dimensions for I222 and the initial P22121 space groups for two of 
>the data sets are:
>I222: a=87.8 b=101.18 c=123.63; P22121: a=93.34 b=105.47 c=122.98;
>
>I superposed the molecule in I222 onto one of the two located for the 
>initially solved P22121; the orientation of the NCS-related molecule in P22121 
>differs from the crystallographic-symmetry related one in I222. Trying to 
>solve this P22121 data set in I222 with MR, does not result in high Z scores, 
>and maps do not look good.
>
>Some of the data sets that process in I222 to ~ 3 Angstr., I can also solve in 
>P22121, locating two molecules (differences may not be that clear in this 
>case, since the resolution is lower).
>
>Some other data sets process in P22121 with Aimless; with a substantial 
>off-origin Patterson peak, indicating translational NCS. For these, Phaser 
>positions two molecules that are related by crystallographic translational 
>NCS. These two molecules are crystallographic-symmetry related in the original 
>P22121 data set. I can also solve these data sets in I222 space group, with 
>the overall Z score higher than for the P22121 data. 
>
>I am uncertain, what the �true� space group for some of my data sets is. Could 
>it be that for data that process in P22121, but can be solved in I222, 
>reflections that would indicate I222 space group were not collected? 
>Alternatively, perhaps what I am seeing is that there is a (gradual) 
>transition of the crystal lattice (between P22121 and I222 or vice versa), 
>caused by variation in crystal handling/cooling or exposure to X-rays.
>
>It�s relevant to me, because in P22121 space group, a region of the molecule 
>that is of biological interest makes NCS-related crystal contacts that are 
>crystallographic-symmetry related in I222.
>
>Has anybody observed similar cases? I would appreciate comments.
>
>Cheers,
>
>Florian
>
>
>

Re: [ccp4bb] I222 - P22121 space-group ambiguity

[Jurgen Bosch]

I think Eleanor was looking at the cell axis in particular a and b they are off 
by almost 10%, hence unlikely to be identical, unless I missed something there.
Jürgen
..
Jürgen Bosch
Johns Hopkins University
Bloomberg School of Public Health
Department of Biochemistry & Molecular Biology
Johns Hopkins Malaria Research Institute
615 North Wolfe Street, W8708
Baltimore, MD 21205
Office: +1-410-614-4742
Lab:  +1-410-614-4894
Fax:  +1-410-955-2926
http://lupo.jhsph.edu

On Oct 13, 2014, at 11:30 AM, Bernhard Rupp 
mailto:b...@ruppweb.org>> wrote:

It might help to look at the images and predictions. You a have serial vs 
integral extinctions
(i.e. conditions limiting reflections are:)
P 2 21 21 (Standard: P 21 21 2, btw)
0K0 : K=2N only
00L : L=2N only

I222
HKL : H+K+L=2N only

Alternatively you could process unmerged data with XPREP or so and look
at the systematic absence violations.

The lattice metric  is the same for both SGs (did not understand Eleanor’s 
remark)

In principle, differences between NCS and XS of compatible metric become more 
distinct at
higher resolution (i.e. at night, all cats are grey).

Best, BR

From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Florian 
Schmitzberger
Sent: Montag, 13. Oktober 2014 10:47
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] I222 - P22121 space-group ambiguity

Hi everybody,

I collected a number of X-ray data sets from crystals originating from the same 
cryst. drop. I solved the initial structure in P22121 space group by MR with 
Phaser locating two molecules (data to ~ 2.1 Angstr.); refined R/Rfree: 
0.213/0.244.

Processing of some of the other data sets with XDS/Aimless is consistent with 
I222 space group (resolution ~ 2.6 Ang.). I can locate one molecule. The 
unit-cell dimensions for I222 and the initial P22121 space groups for two of 
the data sets are:
I222: a=87.8 b=101.18 c=123.63; P22121: a=93.34 b=105.47 c=122.98;

I superposed the molecule in I222 onto one of the two located for the initially 
solved P22121; the orientation of the NCS-related molecule in P22121 differs 
from the crystallographic-symmetry related one in I222. Trying to solve this 
P22121 data set in I222 with MR, does not result in high Z scores, and maps do 
not look good.

Some of the data sets that process in I222 to ~ 3 Angstr., I can also solve in 
P22121, locating two molecules (differences may not be that clear in this case, 
since the resolution is lower).

Some other data sets process in P22121 with Aimless; with a substantial 
off-origin Patterson peak, indicating translational NCS. For these, Phaser 
positions two molecules that are related by crystallographic translational NCS. 
These two molecules are crystallographic-symmetry related in the original 
P22121 data set. I can also solve these data sets in I222 space group, with the 
overall Z score higher than for the P22121 data.

I am uncertain, what the ‘true’ space group for some of my data sets is. Could 
it be that for data that process in P22121, but can be solved in I222, 
reflections that would indicate I222 space group were not collected? 
Alternatively, perhaps what I am seeing is that there is a (gradual) transition 
of the crystal lattice (between P22121 and I222 or vice versa), caused by 
variation in crystal handling/cooling or exposure to X-rays.

It’s relevant to me, because in P22121 space group, a region of the molecule 
that is of biological interest makes NCS-related crystal contacts that are 
crystallographic-symmetry related in I222.

Has anybody observed similar cases? I would appreciate comments.

Cheers,

Florian

Re: [ccp4bb] I222 - P22121 space-group ambiguity

[Harry Powell]

Hi Bernhard et al

> 
> The lattice metric  is the same for both SGs (did not understand Eleanor’s 
> remark)

>> Hmmm - 
>> well the I212121 cell and the P21221 cells you give here have different 
>> volumes, so they cant be just rearrangements of symmetry operators and 
>> non-cryst translation.
>> 


Cell volume for the I222 cell = 87.8 * 101.18 * 123.63 ~= 1,098,280 Å^3

Cell volume for the P22121 cell = 93.34 * 105.47 * 122.98 ~=1,210,685 Å^3

Or ~10% different - so to me, anyway, Eleanor's remark made perfect sense.

>  
> In principle, differences between NCS and XS of compatible metric become more 
> distinct at
> higher resolution (i.e. at night, all cats are grey).
>  
> Best, BR
>  
> From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Florian 
> Schmitzberger
> Sent: Montag, 13. Oktober 2014 10:47
> To: CCP4BB@JISCMAIL.AC.UK
> Subject: [ccp4bb] I222 - P22121 space-group ambiguity
>  
> Hi everybody,
>  
> I collected a number of X-ray data sets from crystals originating from the 
> same cryst. drop. I solved the initial structure in P22121 space group by MR 
> with Phaser locating two molecules (data to ~ 2.1 Angstr.); refined R/Rfree: 
> 0.213/0.244.
>  
> Processing of some of the other data sets with XDS/Aimless is consistent with 
> I222 space group (resolution ~ 2.6 Ang.). I can locate one molecule. The 
> unit-cell dimensions for I222 and the initial P22121 space groups for two of 
> the data sets are:
> I222: a=87.8 b=101.18 c=123.63; P22121: a=93.34 b=105.47 c=122.98;
>  
> I superposed the molecule in I222 onto one of the two located for the 
> initially solved P22121; the orientation of the NCS-related molecule in 
> P22121 differs from the crystallographic-symmetry related one in I222. Trying 
> to solve this P22121 data set in I222 with MR, does not result in high Z 
> scores, and maps do not look good.
>  
> Some of the data sets that process in I222 to ~ 3 Angstr., I can also solve 
> in P22121, locating two molecules (differences may not be that clear in this 
> case, since the resolution is lower).
>  
> Some other data sets process in P22121 with Aimless; with a substantial 
> off-origin Patterson peak, indicating translational NCS. For these, Phaser 
> positions two molecules that are related by crystallographic translational 
> NCS. These two molecules are crystallographic-symmetry related in the 
> original P22121 data set. I can also solve these data sets in I222 space 
> group, with the overall Z score higher than for the P22121 data. 
>  
> I am uncertain, what the ‘true’ space group for some of my data sets is. 
> Could it be that for data that process in P22121, but can be solved in I222, 
> reflections that would indicate I222 space group were not collected? 
> Alternatively, perhaps what I am seeing is that there is a (gradual) 
> transition of the crystal lattice (between P22121 and I222 or vice versa), 
> caused by variation in crystal handling/cooling or exposure to X-rays.
>  
> It’s relevant to me, because in P22121 space group, a region of the molecule 
> that is of biological interest makes NCS-related crystal contacts that are 
> crystallographic-symmetry related in I222.
>  
> Has anybody observed similar cases? I would appreciate comments.
>  
> Cheers,
>  
> Florian
>  
>  

Harry
--
Dr Harry Powell, MRC Laboratory of Molecular Biology, Francis Crick Avenue, 
Cambridge Biomedical Campus, Cambridge CB2 0QH
Chairman of International Union of Crystallography Commission on 
Crystallographic Computing
Chairman of European Crystallographic Association SIG9 (Crystallographic 
Computing) 

Re: [ccp4bb] I222 - P22121 space-group ambiguity

[Isupov, Michail]

Hi Florian,

When your pseudo-translation (native Patterson peak) is close to (0.5,0.5,0.5) 
even in p22121 you will have have most of reflections with
h+k+l=2n+1 
measured as weak. In your lower resolution datasets 2.6 and 3.0 A non-weak 
reflections can be lost in noise and your group assignment
may go wrong, and all of your crystals may be P22121.

Having said this, polymorphism is quite common.
In a recent publication Acta Crystallogr D Biol Crystallogr. 2014 Sep 1;70(Pt 
9):2430-43, Lebedev & Isupov
we describe a case where two crystals had similar cell parameters in different 
space groups

a = 119.2, b = 192.5, c = 77.3 P212121
and
a = 112.0, b = 192.2, c = 76.7 P21212.

Both structures could be refined in the other space group at 1.8A resolution, 
but resulting R-frees were 8-15 % higher in the wrong space group.
Both space group appeared as a result of symmetry breakdown in a supergroup 
A2122 (reindexed C2221).
P212121  structure could also be refined in the supergroup with FreeR only 3 % 
higher than in the real space group.
However as half of reflections had to be thrown out to reindex to A2122 even if 
FreeR would be equal in the supergroup and P212121
I would consider the latter correct.

I would recommend to  solve and refine your low resolution structures in all 
possible space groups and choose the one with  better refinement statistics,
preferring  not centered one in case of equal Rfrees.

Quality of density maps could be criteria in borderline cases.

Cheers,

Misha



From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Florian 
Schmitzberger [schmitzber...@crystal.harvard.edu]
Sent: 13 October 2014 09:47
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] I222 - P22121 space-group ambiguity

Hi everybody,

I collected a number of X-ray data sets from crystals originating from the same 
cryst. drop. I solved the initial structure in P22121 space group by MR with 
Phaser locating two molecules (data to ~ 2.1 Angstr.); refined R/Rfree: 
0.213/0.244.

Processing of some of the other data sets with XDS/Aimless is consistent with 
I222 space group (resolution ~ 2.6 Ang.). I can locate one molecule. The 
unit-cell dimensions for I222 and the initial P22121 space groups for two of 
the data sets are:
I222: a=87.8 b=101.18 c=123.63; P22121: a=93.34 b=105.47 c=122.98;

I superposed the molecule in I222 onto one of the two located for the initially 
solved P22121; the orientation of the NCS-related molecule in P22121 differs 
from the crystallographic-symmetry related one in I222. Trying to solve this 
P22121 data set in I222 with MR, does not result in high Z scores, and maps do 
not look good.

Some of the data sets that process in I222 to ~ 3 Angstr., I can also solve in 
P22121, locating two molecules (differences may not be that clear in this case, 
since the resolution is lower).

Some other data sets process in P22121 with Aimless; with a substantial 
off-origin Patterson peak, indicating translational NCS. For these, Phaser 
positions two molecules that are related by crystallographic translational NCS. 
These two molecules are crystallographic-symmetry related in the original 
P22121 data set. I can also solve these data sets in I222 space group, with the 
overall Z score higher than for the P22121 data.

I am uncertain, what the ‘true’ space group for some of my data sets is. Could 
it be that for data that process in P22121, but can be solved in I222, 
reflections that would indicate I222 space group were not collected? 
Alternatively, perhaps what I am seeing is that there is a (gradual) transition 
of the crystal lattice (between P22121 and I222 or vice versa), caused by 
variation in crystal handling/cooling or exposure to X-rays.

It’s relevant to me, because in P22121 space group, a region of the molecule 
that is of biological interest makes NCS-related crystal contacts that are 
crystallographic-symmetry related in I222.

Has anybody observed similar cases? I would appreciate comments.

Cheers,

Florian

[ccp4bb] Reminder: CCP4 Study Weekend 2015 - Travel bursary application deadline is 31st of October

[Ronan Keegan]

*The CCP4 Study Weekend (7 - 9 January 2015)*
*East Midlands Conference Centre, University of Nottingham*
**
*Wednesday 7 - MX User Meeting*
*Thursday 8 / Friday 9 - CCP4 Study Weekend*
**
*"Advances in Experimental Phasing"*
**
*We cordially invite you to participate in the 2015 CCP4 Study Weekend 
at the the East Midlands Conference Centre, University of Nottingham. 
The annual CCP4 Study Weekend is a chance to shake off the post-New Year 
torpor, and work hard and play hard with your fellow crystallographers. 
Once again, we have put together an exciting scientific programme for 
the Thursday and Friday, either side of the traditional conference 
dinner. Please also check out the satellite meetings which may be of 
interest. The Study Weekend is a chance to catch up with old friends, 
but is also a chance to meet the CCP4 staff who will be there to present 
and demonstrate the latest software and to answer questions - please say 
hello!*

**
*This year, the topic for the Study Weekend is "Advances in Experimental 
Phasing". In keeping with previous CCP4 meetings, the lectures will 
focus on the presentation and discussion of advanced methods and 
techniques developed and used by the leaders in the field.

***
*Scientific Organisers*
*Thomas Schneider - EMBL Hamburg (Germany)*
*Airlie McCoy - University of Cambridge (UK)*

*Further details of the program and the registration are at 
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Re: [ccp4bb] I222 - P22121 space-group ambiguity

[Ed Pozharski]

Yes, having different crystal forms in the same crystallization conditions is, 
while clearly uncommon, not unheard of. I had a case once where at least four 
different crystal forms were observed in crystals harvested from identically 
prepared drops.  It may be that there is some major set of contacts that 
promote certain arrangement of protein molecules (fiber-like), and these can 
then be packed in multiple ways as controlled by another ser of weaker 
interactions. 


Sent on a Sprint Samsung Galaxy S® III

 Original message From: Florian Schmitzberger 
 Date:10/13/2014  4:47 AM  
(GMT-05:00) To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] 
I222 - P22121 space-group ambiguity 
Hi everybody,

I collected a number of X-ray data sets from crystals originating from the same 
cryst. drop. I solved the initial structure in P22121 space group by MR with 
Phaser locating two molecules (data to ~ 2.1 Angstr.); refined R/Rfree: 
0.213/0.244.

Processing of some of the other data sets with XDS/Aimless is consistent with 
I222 space group (resolution ~ 2.6 Ang.). I can locate one molecule. The 
unit-cell dimensions for I222 and the initial P22121 space groups for two of 
the data sets are:
I222: a=87.8 b=101.18 c=123.63; P22121: a=93.34 b=105.47 c=122.98;

I superposed the molecule in I222 onto one of the two located for the initially 
solved P22121; the orientation of the NCS-related molecule in P22121 differs 
from the crystallographic-symmetry related one in I222. Trying to solve this 
P22121 data set in I222 with MR, does not result in high Z scores, and maps do 
not look good.

Some of the data sets that process in I222 to ~ 3 Angstr., I can also solve in 
P22121, locating two molecules (differences may not be that clear in this case, 
since the resolution is lower).

Some other data sets process in P22121 with Aimless; with a substantial 
off-origin Patterson peak, indicating translational NCS. For these, Phaser 
positions two molecules that are related by crystallographic translational NCS. 
These two molecules are crystallographic-symmetry related in the original 
P22121 data set. I can also solve these data sets in I222 space group, with the 
overall Z score higher than for the P22121 data. 

I am uncertain, what the ‘true’ space group for some of my data sets is. Could 
it be that for data that process in P22121, but can be solved in I222, 
reflections that would indicate I222 space group were not collected? 
Alternatively, perhaps what I am seeing is that there is a (gradual) transition 
of the crystal lattice (between P22121 and I222 or vice versa), caused by 
variation in crystal handling/cooling or exposure to X-rays.

It’s relevant to me, because in P22121 space group, a region of the molecule 
that is of biological interest makes NCS-related crystal contacts that are 
crystallographic-symmetry related in I222.

Has anybody observed similar cases? I would appreciate comments.

Cheers,

Florian

Re: [ccp4bb] I222 - P22121 space-group ambiguity

[Bernhard Rupp]

It might help to look at the images and predictions. You a have serial vs
integral extinctions 

(i.e. conditions limiting reflections are:)

P 2 21 21 (Standard: P 21 21 2, btw)

0K0 : K=2N only 

00L : L=2N only  

 

I222

HKL : H+K+L=2N only

 

Alternatively you could process unmerged data with XPREP or so and look

at the systematic absence violations.

 

The lattice metric  is the same for both SGs (did not understand Eleanor's
remark)

 

In principle, differences between NCS and XS of compatible metric become
more distinct at 

higher resolution (i.e. at night, all cats are grey).

 

Best, BR

 

From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of
Florian Schmitzberger
Sent: Montag, 13. Oktober 2014 10:47
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] I222 - P22121 space-group ambiguity

 

Hi everybody,

 

I collected a number of X-ray data sets from crystals originating from the
same cryst. drop. I solved the initial structure in P22121 space group by MR
with Phaser locating two molecules (data to ~ 2.1 Angstr.); refined R/Rfree:
0.213/0.244.

 

Processing of some of the other data sets with XDS/Aimless is consistent
with I222 space group (resolution ~ 2.6 Ang.). I can locate one molecule.
The unit-cell dimensions for I222 and the initial P22121 space groups for
two of the data sets are:

I222: a=87.8 b=101.18 c=123.63; P22121: a=93.34 b=105.47 c=122.98;

 

I superposed the molecule in I222 onto one of the two located for the
initially solved P22121; the orientation of the NCS-related molecule in
P22121 differs from the crystallographic-symmetry related one in I222.
Trying to solve this P22121 data set in I222 with MR, does not result in
high Z scores, and maps do not look good.

 

Some of the data sets that process in I222 to ~ 3 Angstr., I can also solve
in P22121, locating two molecules (differences may not be that clear in this
case, since the resolution is lower).

 

Some other data sets process in P22121 with Aimless; with a substantial
off-origin Patterson peak, indicating translational NCS. For these, Phaser
positions two molecules that are related by crystallographic translational
NCS. These two molecules are crystallographic-symmetry related in the
original P22121 data set. I can also solve these data sets in I222 space
group, with the overall Z score higher than for the P22121 data. 

 

I am uncertain, what the 'true' space group for some of my data sets is.
Could it be that for data that process in P22121, but can be solved in I222,
reflections that would indicate I222 space group were not collected?
Alternatively, perhaps what I am seeing is that there is a (gradual)
transition of the crystal lattice (between P22121 and I222 or vice versa),
caused by variation in crystal handling/cooling or exposure to X-rays.

 

It's relevant to me, because in P22121 space group, a region of the molecule
that is of biological interest makes NCS-related crystal contacts that are
crystallographic-symmetry related in I222.

 

Has anybody observed similar cases? I would appreciate comments.

 

Cheers,

 

Florian

 

 

Re: [ccp4bb] I222 - P22121 space-group ambiguity

[Jodie Johnston]

Hi Florian,



Not sure if this is helpful or not but I have a particular protein that seems 
to be either P22121 (with 2 molecules) or I222 (with one molecule).

Not from the same drop but crystallised in similar conditions but with 
different small molecules bound.

In many cases the crystals can be processed in both space groups but refine 
better in one over the other.

I always thought (as the small molecules affect some loops folding on the 
active site in my case)

that sometimes all my protein molecules had the same conformation of those 
loops and in other cases there were subtle differences - do you see any 
differences or small molecules bound asymmetrically in your P22121 cases ?





Best regards



Jodie


From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Florian 
Schmitzberger [schmitzber...@crystal.harvard.edu]
Sent: Monday, 13 October 2014 9:47 p.m.
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] I222 - P22121 space-group ambiguity

Hi everybody,

I collected a number of X-ray data sets from crystals originating from the same 
cryst. drop. I solved the initial structure in P22121 space group by MR with 
Phaser locating two molecules (data to ~ 2.1 Angstr.); refined R/Rfree: 
0.213/0.244.

Processing of some of the other data sets with XDS/Aimless is consistent with 
I222 space group (resolution ~ 2.6 Ang.). I can locate one molecule. The 
unit-cell dimensions for I222 and the initial P22121 space groups for two of 
the data sets are:
I222: a=87.8 b=101.18 c=123.63; P22121: a=93.34 b=105.47 c=122.98;

I superposed the molecule in I222 onto one of the two located for the initially 
solved P22121; the orientation of the NCS-related molecule in P22121 differs 
from the crystallographic-symmetry related one in I222. Trying to solve this 
P22121 data set in I222 with MR, does not result in high Z scores, and maps do 
not look good.

Some of the data sets that process in I222 to ~ 3 Angstr., I can also solve in 
P22121, locating two molecules (differences may not be that clear in this case, 
since the resolution is lower).

Some other data sets process in P22121 with Aimless; with a substantial 
off-origin Patterson peak, indicating translational NCS. For these, Phaser 
positions two molecules that are related by crystallographic translational NCS. 
These two molecules are crystallographic-symmetry related in the original 
P22121 data set. I can also solve these data sets in I222 space group, with the 
overall Z score higher than for the P22121 data.

I am uncertain, what the ‘true’ space group for some of my data sets is. Could 
it be that for data that process in P22121, but can be solved in I222, 
reflections that would indicate I222 space group were not collected? 
Alternatively, perhaps what I am seeing is that there is a (gradual) transition 
of the crystal lattice (between P22121 and I222 or vice versa), caused by 
variation in crystal handling/cooling or exposure to X-rays.

It’s relevant to me, because in P22121 space group, a region of the molecule 
that is of biological interest makes NCS-related crystal contacts that are 
crystallographic-symmetry related in I222.

Has anybody observed similar cases? I would appreciate comments.

Cheers,

Florian

Re: [ccp4bb] I222 - P22121 space-group ambiguity

[Eleanor Dodson]

Hmmm -
well the I212121 cell and the P21221 cells you give here have different
volumes, so they cant be just rearrangements of symmetry operators and
non-cryst translation.

Do you get other sets of cell dimensions for different processing?

Eleanor

On 13 October 2014 09:47, Florian Schmitzberger <
schmitzber...@crystal.harvard.edu> wrote:

> Hi everybody,
>
> I collected a number of X-ray data sets from crystals originating from the
> same cryst. drop. I solved the initial structure in P22121 space group by
> MR with Phaser locating two molecules (data to ~ 2.1 Angstr.); refined
> R/Rfree: 0.213/0.244.
>
> Processing of some of the other data sets with XDS/Aimless is consistent
> with I222 space group (resolution ~ 2.6 Ang.). I can locate one molecule.
> The unit-cell dimensions for I222 and the initial P22121 space groups for
> two of the data sets are:
> I222: a=87.8 b=101.18 c=123.63; P22121: a=93.34 b=105.47 c=122.98;
>
> I superposed the molecule in I222 onto one of the two located for the
> initially solved P22121; the orientation of the NCS-related molecule in
> P22121 differs from the crystallographic-symmetry related one in I222.
> Trying to solve this P22121 data set in I222 with MR, does not result in
> high Z scores, and maps do not look good.
>
> Some of the data sets that process in I222 to ~ 3 Angstr., I can also
> solve in P22121, locating two molecules (differences may not be that clear
> in this case, since the resolution is lower).
>
> Some other data sets process in P22121 with Aimless; with a substantial
> off-origin Patterson peak, indicating translational NCS. For these, Phaser
> positions two molecules that are related by crystallographic translational
> NCS. These two molecules are crystallographic-symmetry related in the
> original P22121 data set. I can also solve these data sets in I222 space
> group, with the overall Z score higher than for the P22121 data.
>
> I am uncertain, what the ‘true’ space group for some of my data sets is.
> Could it be that for data that process in P22121, but can be solved in
> I222, reflections that would indicate I222 space group were not collected?
> Alternatively, perhaps what I am seeing is that there is a (gradual)
> transition of the crystal lattice (between P22121 and I222 or vice versa),
> caused by variation in crystal handling/cooling or exposure to X-rays.
>
> It’s relevant to me, because in P22121 space group, a region of the
> molecule that is of biological interest makes NCS-related crystal contacts
> that are crystallographic-symmetry related in I222.
>
> Has anybody observed similar cases? I would appreciate comments.
>
> Cheers,
>
> Florian
>
>
>

[ccp4bb] I222 - P22121 space-group ambiguity

[Florian Schmitzberger]

Hi everybody,

I collected a number of X-ray data sets from crystals originating from the same 
cryst. drop. I solved the initial structure in P22121 space group by MR with 
Phaser locating two molecules (data to ~ 2.1 Angstr.); refined R/Rfree: 
0.213/0.244.

Processing of some of the other data sets with XDS/Aimless is consistent with 
I222 space group (resolution ~ 2.6 Ang.). I can locate one molecule. The 
unit-cell dimensions for I222 and the initial P22121 space groups for two of 
the data sets are:
I222: a=87.8 b=101.18 c=123.63; P22121: a=93.34 b=105.47 c=122.98;

I superposed the molecule in I222 onto one of the two located for the initially 
solved P22121; the orientation of the NCS-related molecule in P22121 differs 
from the crystallographic-symmetry related one in I222. Trying to solve this 
P22121 data set in I222 with MR, does not result in high Z scores, and maps do 
not look good.

Some of the data sets that process in I222 to ~ 3 Angstr., I can also solve in 
P22121, locating two molecules (differences may not be that clear in this case, 
since the resolution is lower).

Some other data sets process in P22121 with Aimless; with a substantial 
off-origin Patterson peak, indicating translational NCS. For these, Phaser 
positions two molecules that are related by crystallographic translational NCS. 
These two molecules are crystallographic-symmetry related in the original 
P22121 data set. I can also solve these data sets in I222 space group, with the 
overall Z score higher than for the P22121 data. 

I am uncertain, what the ‘true’ space group for some of my data sets is. Could 
it be that for data that process in P22121, but can be solved in I222, 
reflections that would indicate I222 space group were not collected? 
Alternatively, perhaps what I am seeing is that there is a (gradual) transition 
of the crystal lattice (between P22121 and I222 or vice versa), caused by 
variation in crystal handling/cooling or exposure to X-rays.

It’s relevant to me, because in P22121 space group, a region of the molecule 
that is of biological interest makes NCS-related crystal contacts that are 
crystallographic-symmetry related in I222.

Has anybody observed similar cases? I would appreciate comments.

Cheers,

Florian