Re: [ccp4bb] low resolution map with unmodelled map

[Roger Rowlett]

I would back way up in the process and verify that the basics are correct.
Ten molecules in the ASU is unusual unless your molecules are forming
defined oligomers, e.g., 2.5 tetramers, etc. Precise predictions of
monomers in the ASU by Matthews number is increasingly unreliable above 2-3
monomers. You have to look at lattice packing to be more sure of the
correct number. If there are defined oligomers, e.g.dimers, the initial
search may work better with oligomers as the search unit than monomers.

How confident are you of the space group and unit cell assignment? What
does the lattice packing of the MR solution look like? (You can do this in
Coot.) Do protein molecules in the lattice make sensible contacts? Are
there serious overlaps? Can you see solvent channels? Can you see
symmetry-related oligomers? I'd sort this out before proceeding further. If
these check out, then there are ways of using noncrystallographic symmetry
in your ASU to significantly improve your initial maps for rebuilding,
although sequence assignment may still be challenging at this resolution.

Roger Rowlett


On Mon, Aug 19, 2019, 8:15 PM Zhu Qiao  wrote:

> Sorry for the initial message. I tried to attach Matthews coefficient
> calculation figure but failed to do so, which resulted the message as not
> plain text.  Below is my question, thanks.
>
> I collected one dataset and processed it to 3.6 angstrom. my protein is
> quite small with only 14 kDa. It is estimated over ten molecules in one ASU
> based on Matthew coefficient calculation.
>
> However, only ten molecules can be correctly placed with good fitting. I
> can observe extra Fo-Fc electron density maps there needed to be modelled.
> But Fo-Fc electron density maps are discontinuous.
>
> I tried to fix the ten molecules as the partial solution in phaser and
> search more molecules, either resulted in no solution or the newly added
> molecules didn't fit in the map after refinement.
>
> So I manually built the poly alanine chain in order to decrease the R
> factor. I built around 200 amino acids into the final model. but these poly
> alanine model can hardly be interpreted to remodel as my target protein
> because of the low resolution. Currently the ten molecules model has a
> Rwork/free as 0.33/0.36. The model with poly alanine chain has a
> Rwork/free as 0.30/0.34.  I can already extract the useful information
> based on the well fitted ten molecules. And the fitting of the poly alanine
> models could just for better model refinement.
>
> Can I get some suggestions regarding this kind of issue, what's the
> general practise for such situation. Can I deposit the model with poly
> alanine fitted and labelled it unidentified to pdb?
>
> Thanks for any suggestions and replies.
>
> Best Regards
> Qiao Zhu
>
> 
>
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[ccp4bb] low resolution map with unmodelled map

[Zhu Qiao]

Sorry for the initial message. I tried to attach Matthews coefficient 
calculation figure but failed to do so, which resulted the message as not plain 
text.  Below is my question, thanks. 

I collected one dataset and processed it to 3.6 angstrom. my protein is quite 
small with only 14 kDa. It is estimated over ten molecules in one ASU based on 
Matthew coefficient calculation.

However, only ten molecules can be correctly placed with good fitting. I can 
observe extra Fo-Fc electron density maps there needed to be modelled. But 
Fo-Fc electron density maps are discontinuous. 

I tried to fix the ten molecules as the partial solution in phaser and search 
more molecules, either resulted in no solution or the newly added molecules 
didn't fit in the map after refinement. 

So I manually built the poly alanine chain in order to decrease the R factor. I 
built around 200 amino acids into the final model. but these poly alanine model 
can hardly be interpreted to remodel as my target protein because of the low 
resolution. Currently the ten molecules model has a Rwork/free as 0.33/0.36. 
The model with poly alanine chain has a  Rwork/free as 0.30/0.34.  I can 
already extract the useful information based on the well fitted ten molecules. 
And the fitting of the poly alanine models could just for better model 
refinement. 

Can I get some suggestions regarding this kind of issue, what's the general 
practise for such situation. Can I deposit the model with poly alanine fitted 
and labelled it unidentified to pdb?  

Thanks for any suggestions and replies. 

Best Regards 
Qiao Zhu



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[ccp4bb] low resolution map with unmodeled maps

[Zhu Qiao]

Dear all

I collected one dataset and processed it to 3.6 angstrom. my protein is quite 
small with only 14 kDa. It is estimated over ten molecules in one ASU based on 
Matthew coefficient calculation.

However, only ten molecules can be correctly placed with good fitting. I can 
observe extra Fo-Fc electron density maps there needed to be modelled. But 
Fo-Fc electron density maps are discontinuous. 

I tried to fix the ten molecules as the partial solution in phaser and search 
more molecules, either resulted in no solution or the newly added molecules 
didn't fit in the map after refinement. 

So I manually built the poly alanine chain in order to decrease the R factor. I 
built around 200 amino acids into the final model. but these poly alanine model 
can hardly be interpreted to remodel as my target protein because of the low 
resolution. Currently the ten molecules model has a Rwork/free as 0.33/0.36. 
The model with poly alanine chain has a  Rwork/free as 0.30/0.34.  I can 
already extract the useful information based on the well fitted ten molecules. 
And the fitting of the poly alanine models could just for better model 
refinement. 

Can I get some suggestions regarding this kind of issue, what's the general 
practise for such situation. Can I deposit the model with poly alanine fitted 
and labelled it unidentified to pdb?  

Thanks for any suggestions and replies. 

Best Regards 
Qiao Zhu



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[ccp4bb] Multiple Postdoctoral Fellowships available at the “Multiscale Research Institute for Complex Systems” at Fudan University of Shanghai

[lyguo]

Dear All,
The Multiscale Research Institute for Complex Systems (MRICS) at Fudan 
University is located at the Zhangjiang Campus of Fudan University and is 
supported by the Shanghai High-level Talents Program.MRICS is strongly 
committed to the development of noveland effective multi-scale imaging 
technology that spans microscopic molecular structures all the way to 
macroscopic medical imaging, with the aim to provide unprecedented spatial and 
temporal insights into the structures and functions of living beings at all 
levels (molecules, cells, tissues, organs and even whole organisms). 
Specifically for structural biology, MRICS isequipped witha state-of-the-art 
cryo-EM facility that includesFEI Titan Krios equipped with Volta phase plate, 
Glacios, Talos and Aquilos. MRICS is also located next to Shanghai Synchrotron 
Radiation Facility for X-ray crystallography.
Our team includes Nobel laureate and international leading interdisciplinary 
experts. 
We have new openings for multiple postdoctoral fellowsinstructural biologywho 
will be mainly involved in structural studies of important biological systems 
by means of X-ray crystallography and cryo-electron microscopy including 
single-particle and tomography.
Requirements:
The applicants should have a recent Ph.D. degree(within three years of 
graduation)or will have a Ph.D. degree within the next six months in biology or 
chemistry-related fields, aredevoted to excellence in scientific research, have 
strong sense of responsibility, and arehighly motivated andhardworking. For 
these positions,extensive experience in protein expression and purification is 
a must, while prior experience in X-ray crystallography or cryo-EM is a plus, 
but is not required. 
Compensation:
1) We offer internationally competitive salary and fringe benefits, the level 
of salary will be determined according to the applicant's experience and 
qualification;
2) We will assist in applying for low-rent housing in Shanghai and provide 
certain housing subsidies;
3) We have ample opportunities to collaborate with renown laboratories 
worldwide;
4) We provide support for applying for funding opportunities whenever 
applicable. 
Shanghai is one of the most cosmopolitan cities in China with strong economy 
and vibrant scientific community.
For interested applicants, please submit postdoctoral application packages (a 
combined pdf) including resumes, concise research interest statements, 
representative publications, phone numbers and email addresses of three 
academic referees to:mrics...@fudan.edu.cn
We look forward to your joining of our first-class team!



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Re: [ccp4bb] ORCID

[Crissy L. Tarver]

Hi Joe,

I recently submitted 4 structures to the PDB, and I found that an ORCID was
necessary for the corresponding author. However, additional authors were
added without an ORCID.

Hope this is somewhat helpful.

Best,
Crissy


On Mon, Aug 19, 2019 at 2:55 PM Jie Liu  wrote:

> Dear all,
>
> It's been a while since last time I deposited structures to PDB. Do I
> really need an ORCID (Open Researcher and Contributor IDentifier) now to
> submit files? Is it mandatory?
>
> Thank you!
> Jie
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
>
-- 
Crissy L. Tarver, Doctoral Candidate
Biotechnology Science & Engineering
Department of Biological Sciences
University of Alabama Huntsville

Success is not the key to happiness. Happiness is the key to success. If
you love what you are doing, you will be successful.- Albert Schweitzer



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Re: [ccp4bb] ORCID

[Wim Burmeister]

Hello, 
like a lot of items in the pdb entry, the entry is not mandatory. But using the 
ORCID is a good idea in order to be able to claim easily your work if you have 
a very common name and it may be difficult to find your authorship 
unambigously. 
Best 
Wim 


De: "Jie Liu"  
À: "CCP4BB"  
Envoyé: Lundi 19 Août 2019 21:54:52 
Objet: [ccp4bb] ORCID 

Dear all, 

It's been a while since last time I deposited structures to PDB. Do I really 
need an ORCID (Open Researcher and Contributor IDentifier) now to submit files? 
Is it mandatory? 

Thank you! 
Jie 




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Re: [ccp4bb] ORCID

[John Berrisford]

Dear Jie

 

ORCiD is mandatory for all authors that we communicate with during
deposition (contact authors).  One of the reasons is to allow us ensure that
we are able to associate contact authors with all of their depositions even
when their email address changes. 

These contact details are kept private and are not released into the PDB
archive.

 

ORCiD's are not required for authors who are just listed as entry or
citation authors. But we do encourage ORCiD's to be provided for these
authors if they are available. These author names, and any associated
ORCiD's, are made public upon PDB release. 

 

Please see

https://www.wwpdb.org/documentation/policy#toc_authorship

for further details of the different types of authors in a PDB entry. 

 

I hope this helps

 

Regards

 

John

 

From: CCP4 bulletin board  On Behalf Of Jie Liu
Sent: 19 August 2019 20:55
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] ORCID

 

Dear all,


It's been a while since last time I deposited structures to PDB. Do I really
need an ORCID (Open Researcher and Contributor IDentifier) now to submit
files? Is it mandatory?

Thank you!
Jie

 

  _  

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 =1 




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[ccp4bb] ORCID

[Jie Liu]

Dear all,

It's been a while since last time I deposited structures to PDB. Do I really 
need an ORCID (Open Researcher and Contributor IDentifier) now to submit files? 
Is it mandatory?

Thank you!
Jie



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[ccp4bb] OT: HELIX and SHEET records not read by Pymol

[Chris Fage]

Dear All,

Pymol seems to ignore the HELIX and SHEET records in my coordinate file,
and instead assigns secondary structural features according to the default
method. Unless my understanding is flawed, these records should override
Pymol's assignment...?

I generated the records with the Stride Web Interface and converted them to
PDB format with Emma Rath's website (
http://www.canoz.com/sdh/STRIDEtoPDBsecondarystruct.pl).

Does anyone have any ideas? Thanks!

Best wishes,
Chris


Virus-free.
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Re: [ccp4bb] Better Beamline suggestion!

[Andreas Förster]

Dear Chandra,

where at APS did you collect?  NE-CAT (and in particular beamline 24-ID-E)
is well set-up for long unit cells.  You could ask Yury Polikanov (
https://bios.uic.edu/profiles/polikanov-yury/) for advice.  He crystallizes
ribosomes with unit cell that are not quite as big as what you have, but
quite big nevertheless.

An alternative would be P14 at EMBL.  They have measured crystals with unit
cell dimensions beyond 1 kÅ.

All best.


Andreas



On Mon, Aug 19, 2019 at 5:08 PM Chandramohan Kattamuri <
1c5b7cb6c764-dmarc-requ...@jiscmail.ac.uk> wrote:

>
> Dear All
>
> We recently collected a data set at APS, Chicago with unit cell dimensions
> of 68.4; 68.4 and 991.6 A. Our diffraction data extends to 3A with the APS
> set up, however, the long axis has been problematic, resulting in streaking
> of the diffraction data and requires a very specific orientation of the
> crystal for usable diffraction. Can anyone recommend beamlines that can
> give us higher resolution, or a source with a better goniometer allowing
> for more angle manipulation after looping?
>
> Thank a lot
> Chandra K
>
>
>
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
>


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Phone: +41 56 500 21 00 | Direct: +41 56 500 21 76 | Email:
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Re: [ccp4bb] [EXTERNAL] Re: [ccp4bb] Better Beamline suggestion!

[Edward A. Berry]

 I would respectfully suggest that higher pixel resolution does not generally help much 
in these situations. If an average spot is 10 or more pixels wide then the profile is 
defined pretty well. But if the spots overlap, they still overlap with higher pixel 
density. It may make profile fitting more accurate, allowing more accurate 
"deconvolution" of the compound spot into its components, but it will not 
improve the overlap. Smaller or better-focused (on the detector, not the crystal?) beam, 
and longer camera length can help.
  It is analogous to chromatography- If two peaks coming off the column 
overlap, collecting smaller fractions will not help to resolve them. It may 
allow a better-informed decision on the cut-off points when you pool the 
fractions, but it won't separate the overlap.
  On the other hand a multi-circle goniometer is very useful. I remember in one 
of our last trips at SSRL (2007-8?) we used a (Huber 4-circle?) and it was very 
easy to have the long axis in the plane of the image throughout the rotation.  
In the absence of such you can resort to carefully bending the loop or bending 
the pin (Jim Holton made a nifty device for bending the pin) while keeping the 
xtal bathed in the cold stream.


On 08/19/2019 11:12 AM, graeme.win...@diamond.ac.uk wrote:

Chandra

What you are looking for here is a beamline with a detector with many pixels 
(so you can resolve the long axis) and a multi-axis goniometer - probably a 
SmarGon / kappa and an Eiger 16M would make a good combination for this. 
Searching on

https://urldefense.proofpoint.com/v2/url?u=http-3A__biosync.sbkb.org_=DwIFAg=ogn2iPkgF7TkVSicOVBfKg=cFgyH4s-peZ6Pfyh0zB379rxK2XG5oHu7VblrALfYPA=VcmIp54F7yM1JdiEMdBdR0y7xinGb-nsn2-3LI_BHto=5VFyqMBHljO-AEcXr3-pqjF8xFyEejXetVFOxOXLp_Y=

Should allow you to make up a short list

Best wishes Graeme

On 19 Aug 2019, at 16:08, Chandramohan Kattamuri 
<1c5b7cb6c764-dmarc-requ...@jiscmail.ac.uk>
 wrote:


Dear All
We recently collected a data set at APS, Chicago with unit cell dimensions of 
68.4; 68.4 and 991.6 A. Our diffraction data extends to 3A with the APS set up, 
however, the long axis has been problematic, resulting in streaking of the 
diffraction data and requires a very specific orientation of the crystal for 
usable diffraction. Can anyone recommend beamlines that can give us higher 
resolution, or a source with a better goniometer allowing for more angle 
manipulation after looping?
Thank a lot
Chandra K






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Re: [ccp4bb] Better Beamline suggestion!

[graeme.win...@diamond.ac.uk]

Chandra

What you are looking for here is a beamline with a detector with many pixels 
(so you can resolve the long axis) and a multi-axis goniometer - probably a 
SmarGon / kappa and an Eiger 16M would make a good combination for this. 
Searching on

http://biosync.sbkb.org/

Should allow you to make up a short list

Best wishes Graeme

On 19 Aug 2019, at 16:08, Chandramohan Kattamuri 
<1c5b7cb6c764-dmarc-requ...@jiscmail.ac.uk>
 wrote:


Dear All
We recently collected a data set at APS, Chicago with unit cell dimensions of 
68.4; 68.4 and 991.6 A. Our diffraction data extends to 3A with the APS set up, 
however, the long axis has been problematic, resulting in streaking of the 
diffraction data and requires a very specific orientation of the crystal for 
usable diffraction. Can anyone recommend beamlines that can give us higher 
resolution, or a source with a better goniometer allowing for more angle 
manipulation after looping?
Thank a lot
Chandra K






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[ccp4bb] Better Beamline suggestion!

[Chandramohan Kattamuri]

Dear All
We recently collected a data set at APS, Chicago with unit cell dimensions of 
68.4; 68.4 and 991.6 A. Our diffraction data extends to 3A with the APS set up, 
however, the long axis has been problematic, resulting in streaking of the 
diffraction data and requires a very specific orientation of the crystal for 
usable diffraction. Can anyone recommend beamlines that can give us higher 
resolution, or a source with a better goniometer allowing for more angle 
manipulation after looping?

Thank a lot

Chandra K






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[ccp4bb] Postdoc Position in Cryo-EM at The Pennsylvania State University College of Medicine

[Kenneth Lee]

Dear Colleagues,

An exciting research opportunity is available in my team:

Postdoctoral position in Membrane Protein Structural Biology and Biophysics at 
The Pennsylvania State University College of Medicine

The laboratory of Dr. Kenneth Lee seeks applications for a postdoctoral 
position applying cryo-EM approaches to study the structure and mechanism of 
macromolecular devices in biological membranes connected to human disease.  We 
are seeking highly motivated individuals with a PhD or MD/PhD degree in the 
fields of biochemistry, biophysics, neuroscience and molecular cell biology. 
Required is a track-record of productivity. Candidates are expected to have a 
strong work ethic, excellent organizational and communication skills, critical 
thinking abilities and be capable of thriving in a fast-paced environment. The 
ideal candidate will have a strong background in structural biology, molecular 
biology and protein purification. Expertise in membrane protein biochemistry, 
single-particle cryo-EM and/or X-ray crystallography is preferred but not 
required. The successful candidate will join a vibrant community of scientists 
and health professionals and will have on-site access to superb resources 
including a newly established state-of-the-art cryo-EM facility housing a 300 
kV Titan Krios G3i cryo-TEM equipped with a Volta phase plate, Gatan BioQuantum 
energy filter and K3 direct electron detector. Penn State College of Medicine 
is an outstanding environment for researchers investigating molecular 
mechanisms of disease and has a Postdoctoral Affairs Office that provides 
exceptional infrastructure for training and career development. Applicants 
should submit a cover letter, CV and contact information of three references 
to: kenneth...@psu.edu. For more information, visit: https://psu.jobs/job/89930


Sincerely,

Kenneth Lee

Kenneth P.K. Lee, Ph.D.
Assistant Professor
Department of Cellular and Molecular Physiology
Penn State College of Medicine
500 University Drive
Hershey, PA  17033
kenneth...@psu.edu



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