Re: [ccp4bb] number of frames to get a full dataset?

[David Schuller]

The old saying was degrees, not frames. If your frame width is not 1 
degree, the result will differ accordingly.


One factor is whether the detector is centered or offset, and whether it 
is large enough to get the entire pattern. If the detector is offset, 
you are not getting the full diffraction from that position.


I don't think radiation damage figures into this. It might make it more 
difficult to get N degrees, but it does not change how much diffraction 
is necessary.




On 2020-06-22 18:03, Murpholino Peligro wrote:

Hi.
Quick question...
I have seen *somewhere* that to get a 'full dataset we need to collect 
n frames':

at least 180 frames if symmetry is X
at least 90 frames if symmetry is Y
at least 45 frames if symmetry is Z
Can somebody point where is *somewhere*?

...also...
what other factors can change n... besides symmetry and radiation damage?

Thanks



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Re: [ccp4bb] number of frames to get a full dataset?

[Diana Tomchick]

There are lots of places where you could find this information (many textbooks, 
articles, etc.) but one that I use for classes is quite good due to ease of 
understanding. It’s part of the Proceedings of the CCP4 Study Weekend on Data 
Collection and Processing. There are other quite excellent articles in that 
issue, and all are Open Access.

Dauter, Z. (1999) “Data-collection strategies” Acta Cryst. D55, 1703-1717.

https://journals.iucr.org/d/issues/1999/10/00/ba0020/index.html

Diana

**
Diana R. Tomchick
Professor
Departments of Biophysics and Biochemistry
UT Southwestern Medical Center
5323 Harry Hines Blvd.
Rm. ND10.214A
Dallas, TX 75390-8816
diana.tomch...@utsouthwestern.edu
(214) 645-6383 (phone)
(214) 645-6353 (fax)

On Jun 22, 2020, at 5:03 PM, Murpholino Peligro 
mailto:murpholi...@gmail.com>> wrote:


EXTERNAL MAIL

Hi.
Quick question...
I have seen *somewhere* that to get a 'full dataset we need to collect n 
frames':
at least 180 frames if symmetry is X
at least 90 frames if symmetry is Y
at least 45 frames if symmetry is Z
Can somebody point where is *somewhere*?

...also...
what other factors can change n... besides symmetry and radiation damage?

Thanks



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CAUTION: This email originated from outside UTSW. Please be cautious of links 
or attachments, and validate the sender's email address before replying.





UT Southwestern

Medical Center

The future of medicine, today.



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Re: [ccp4bb] number of frames to get a full dataset?

[Srivastava, Dhiraj]

Well, its not no. of frames but minimum degree of crystal rotation. A goggle 
search gave me this article.

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5557013/


Dhiraj

From: CCP4 bulletin board  on behalf of Murpholino 
Peligro 
Sent: Monday, June 22, 2020 5:03 PM
To: CCP4BB@JISCMAIL.AC.UK 
Subject: [ccp4bb] number of frames to get a full dataset?

Hi.
Quick question...
I have seen *somewhere* that to get a 'full dataset we need to collect n 
frames':
at least 180 frames if symmetry is X
at least 90 frames if symmetry is Y
at least 45 frames if symmetry is Z
Can somebody point where is *somewhere*?

...also...
what other factors can change n... besides symmetry and radiation damage?

Thanks



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https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1



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[ccp4bb] number of frames to get a full dataset?

[Murpholino Peligro]

Hi.
Quick question...
I have seen *somewhere* that to get a 'full dataset we need to collect n
frames':
at least 180 frames if symmetry is X
at least 90 frames if symmetry is Y
at least 45 frames if symmetry is Z
Can somebody point where is *somewhere*?

...also...
what other factors can change n... besides symmetry and radiation damage?

Thanks



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Re: [ccp4bb] Thanks and outcome of ([ccp4bb] Looking for method to find similar oligomeric arrangement)

[Jose Duarte]

A further note on assembly search: since April the RCSB PDB offers
structure search for assemblies. See the "Structure Similarity" section in
https://www.rcsb.org/search/advanced. This works only for PDB-deposited
structures but we are working on making it available for 3D coordinates
submitted by users. The search is extremely fast (below a second) and it
will match the assembly globally. Additionally the search results can be
further filtered by other criteria such as resolution or organism.

The search is also available at Structure Summary Pages (e.g.
https://www.rcsb.org/structure/4HHB)  with the "Find Similar Assemblies"
link below the symmetry information on top left.

Jose


On Sun, 21 Jun 2020 at 07:58, Andrew Lovering  wrote:

> Dear All
>
>
> Many thanks to Johann for the suggestion of Topsearch - rather pleasingly
> our tetramer is novel in the tetrameric arrangement, but each of the dimers
> has its DNA recognition helices in agreement with several hits a win
> all round
>
>
> Bringing joy to my lockdown.
>
>
> Best
>
> Andy
>
> --
>
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Re: [ccp4bb] AW: [ccp4bb] Cold cabinet recommendations for AKTA Pure

[Alan Cheung]

Thanks everyone for recommendations.  The AKTA currently lives in a cold
room hence all of our purifications have been optimised at 4C, and i'm
reluctant to transition them to RT.  Also, most of our columns are not
jacketed.







On Sun, 21 Jun 2020 at 18:19, Boniecki, Michal 
wrote:

> Hi Alan,
> I think new AKTA collector can be refrigerated. If this is not an option,
> consider putting only collector in the cold cabinet and cooling down the
> buffers. Also, you can put only columns and fraction collector inside
> cabinet which has side port for such use. Although you will have to use
> small diameter out tubing to avoid sample dilution when use like this.
> System live will be shorter inside cold cabinet due to increased humidity.
>
> Michal
>
> On Jun 21, 2020, at 11:10, Hughes, Jonathan <
> jon.hug...@bot3.bio.uni-giessen.de> wrote:
>
> 
> CAUTION: This email originated from outside of the University of
> Saskatchewan. Do not click links or open attachments unless you recognize
> the sender and know the content is safe. If in doubt, please forward
> suspicious emails to phish...@usask.ca
>
> hi alan,
>
> why do you want to put the thing in a cold cabinet? if you have protease
> problems, there are better ways to get rid of them than exploiting Q10. on
> the other hand, you'll increase viscosity and thus slow down the flow
> rates. also, the high humidity will reduce the lifetime of your Äkta
> dramatically (ours has been running almost continuously for 20 years at
> room temperature and is still going strong [thanks to tina!]). if you
> really to need to keep the sample at 4 °C, you can put the buffer(s) on ice
> and cool the column with a jacket.
>
> best
>
> jon
>
>
>
> *Von:* CCP4 bulletin board  *Im Auftrag von *Paula
> Salgado
> *Gesendet:* Sonntag, 21. Juni 2020 18:50
> *An:* CCP4BB@JISCMAIL.AC.UK
> *Betreff:* Re: [ccp4bb] Cold cabinet recommendations for AKTA Pure
>
>
>
> Hi Alan
>
>
>
> We got one a few years back, from Fisher, although I think Thermo are the
> manufacturer's:
>
>
>
> Refrigerator FRCR4504W Forma chromatography THERMO SCIENTIFIC FORMA
> FRCR4504W
>
>
>
> They also arranged for a heavy metal shelf to hold the AKTA as the normal
> shelves won't hold the weight.
>
>
>
> We only have one AKTA Pure there, with a fraction collector and the rest
> of the cabinet holds all the columns.
>
>
>
> Hope that helps
>
> Paula
>
>
>
>
>
> ===
>
> Dr Paula S. Salgado
> Senior Lecturer in Macromolecular Crystallography
>
> Molecular Mechanisms of Life Lead
>
> *During the current COVID-19 crisis, my working hours tend to focus on
> early afternoons and evenings, so please bear with me if my reply is
> delayed and if I email outside your normal working hours. In any case, I do
> not expect a response outside those hours. Stay home if you're not involved
> in essential work, stay safe.*
>
>
>
>
>
> Newcastle University Biosciences Institute
>
> Faculty of Medical Sciences
> 2nd Floor Cookson Building
> Newcastle University
> Newcastle upon Tyne, NE2 4HH, UK
>
> Tel: +44 (0)191 208 7432
> Email: paula.salg...@ncl.ac.uk
>
>
>
>
> --
>
> *From:* CCP4 bulletin board  on behalf of Alan
> Cheung 
> *Sent:* 21 June 2020 08:12
> *To:* CCP4BB@JISCMAIL.AC.UK 
> *Subject:* [ccp4bb] Cold cabinet recommendations for AKTA Pure
>
>
>
> ⚠ External sender. Take care when opening links or attachments. Do not
> provide your login details.
>
> Dear All
>
>
>
> Apologies for being wildly off topic, but we're trying to find a decent
> cold cabinet for 2x AKTA Pure + F9C + sample pump.  Does anyone have any
> recommendations, especially for sliding door models?
>
>
>
> Cheers,
>
> Alan
>
>
> --
>
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[ccp4bb] Postgraduate Fellowship, Membrane Structural and Functional Biology, Caffrey Lab, Trinity College Dublin, Ireland

[Martin Caffrey]

*Postgraduate Research Fellow Position*

*Membrane Structural and Functional Biology – Caffrey Lab*

*Trinity College Dublin, Ireland*

*Post Summary*.  The Membrane Structural and Functional Biology group seeks
to solve the structure of medically important membrane proteins for use in
structure/function studies and for structure-based therapeutics
development. Target proteins play a role in atherogenesis, cancer,
allergies and pain and in communicable and other non-communicable diseases.
In support of this activity, we seek a highly self-motivated and committed
Postgraduate Research Fellow.

The Fellowship is available for work in the areas of bacterial *quorum
sensing* and *cell coat synthesis*. Many pathogenic bacteria have the
ability to sense when enough of them occupy a particular space to begin
producing a stable biofilm that supports further growth and virulence.
*Pseudomonas
aeruginosa* is one such opportunistic pathogen that engages in this quorum
sensing process. It is responsible for illness and death in individuals
whose immunity has been compromised such as cystic fibrosis and burns
patients. Projects are underway to solve the atomic-resolution structure of
a number of membrane proteins from *P. aeruginosa *and other pathogenic
bacteria involved in cell quorum sensing and cell coat synthesis. The goal
is to use the structural information to understand how these processes take
place at a molecular level and to assist in the design of new and improved
antibiotics and vaccines to counter the global threat of antimicrobial
infections and resistance.

Candidates should have a high-level honours degree or its equivalent in
biochemistry, biophysics, biotechnology, chemistry, immunology,
microbiology, molecular biology, molecular medicine or a related
discipline.

*Application Procedure. *Candidates are asked to submit a letter detailing
education, training and why they feel they are suitable for the position in
an interdisciplinary group. The letter must be accompanied by a full CV to
include the names and contact details of two referees (email addresses and
phone numbers to be provided).

*Closing Date:*  July 10, 2020.  *Expected Start Date:*  September, 2020

*Lab Website* *https://www.tcd.ie/Biochemistry/research/caffrey/
 *

*Applications should be emailed to: **msfbpg2...@gmail.com
*

-- 
Martin Caffrey  martin.caff...@tcd.ie
Schools of Medicine and Biochemistry & Immunology
Trinity Biomedical Sciences Institute, Trinity College Dublin
Room 5.62,152-160 Pearse St., Dublin 2, D02 R590, Ireland
website: http://www.tcd.ie/Biochemistry/research/caffrey/
phone: +353-(0)1-896-4253; mobile: +353-(0)86-818-7704
Skype: martincaffrey



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

[lyguo]

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 novel and 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 
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Specifically for structural biology, MRICS is equipped with a state-of-the-art 
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Radiation Facility for X-ray crystallography.
 
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who will mainly study important biological systems by means of cryo-electron 
microscopy including single-particle and tomography. 
   
Requirements: 
 
The applicants should have a recent Ph.D. degree (within two years of 
graduation) or will have a Ph.D. degree within the next six months in biology 
or chemistry-related fields, be devoted to excellence in scientific research, 
have strong sense of responsibility, and be highly motivated and hardworking.  
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, fringe benefits and yearly 
bonus.  The level of salary will be determined according to the applicant's 
experience and qualification; 
2)Low-rent housing on campus is provided; 
3)There are ample opportunities to collaborate with world-renown 
laboratories;
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, 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] Ideas for improving refinement with anisotopic data.

[vincent Chaptal]

Dear Matthew,

at your resolution, 1.7A, you are seeing a large difference in 
resolution limits. I'd be curious to know how much "anisotropy" you have 
in the single number anisoB. Could you share it?
As to building felxible loops or peptide, I suspect you anisoB to be 
mild (<40A2 for your resolution) and therefore the effect on map 
artefacts to also be mild. Including the high res data is definitely 
your best shot at getting better maps.


Good luck with your modeling.
Vincent

Le 21/06/2020 à 19:11, Gerard Bricogne a écrit :

Dear Matthew,

  You seem to have been unlucky with this data collection. STARANISO
would be prepared to see significant data extending to 2.0A along a*, 2.7A
along b*, and 1.7A along c*. The latter limit, however, involves some guess
work via the fitting by an ellipsoid of a cut-off surface that is very much
perturbed by the fact that you have a both a cusp and a truncation by the
detector edges in that direction. Without those, it looks quite probable
that you would have good solid data extending beyond 1.6A - if only you
could redo the experiment in such a way as to actually measure them ;-) .

  As Eleanor said, it is a bad idea to make decisions about data on the
basis of the "downstream" criteria you were mentioning. STARANISO is not
just a numerical program that estimates numbers and suggests some sensible
actions that it will carry out for you: it is also a 3D visualisation tool
that shows you much more about your data and its possible shortcomings than
any Table1-style numbers ever will. If there are indeed deficiencies in the
data, like here, these 3D pictures will often point out what shortcomings in
your experiment are responsible for them. In that case, a new experiment, if
feasible, is always better than any surgery on the deficient data.

  Please feel free to get in touch off-line if you would like more
details, such as patterns of outliers that we have observed.


  With best wishes,

  The STARANISO developers.

--
On Sun, Jun 21, 2020 at 05:33:53PM +0100, Eleanor Dodson wrote:

*Please* dont throw good 1.8A data for the sake of statistics!
You should see more detail along certain directions
You will publish your structure providing honest details of the anisotropy
(I hope..) but it is the map quality that matters ..
Eleanor

On Sun, 21 Jun 2020 at 15:16, Matthew Snee <
matthew.s...@postgrad.manchester.ac.uk> wrote:


Hi everyone.

I have an x ray structure that I am finishing up, and there are a few
ambiguous regions where the peptide is poorly resolved.

The data is highly anisotrophic, and requires truncation to around 2.4A to
achieve acceptable merging stats, although there is data in the "good"
direction going as high as 1.8-1.9A (determined by merging stats when using
elliptical cutoff).

I have tried feeding my integration outputs to STARANISO with both the
elliptical and spherical cutoffs, but neither produce better results than
Xia2 Dials, as both need to be truncated further than 2.3A before they give
the same refinement stats (i.e it's best to just let refmac/phenix.refine
deal with the anistropy).

As I understand it, anisotrophy can lead to loss of high resolution detail
because weak observations from the high res shells in the bad direction are
down-weighted, So any tricks to improve map quality (or conversely refining
data with poor completeness) would be appreciated.

I'm not holding out hope that I can deposit anything better than 2.3A, but
Improving the maps might really help me with some of these troublesome
loops :)

Cheers

Matthew.

Sent from Outlook Mobile 

--

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--

Vincent Chaptal, PhD

Director of GdR APPICOM

Drug Resistance and Membrane Proteins Lab


MMSB -UMR5086

7 passage du Vercors

69007 LYON

FRANCE

+33 4 37 65 29 01

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