Re: [ccp4bb] need help with background and over fitting in 2D classification

[Schara Safarian]

Hey Suchi,

you could firstly increase the binning to factor to 4-5. Also, it would help to 
shrink the mask radius. Most definitely keep the resolution limited to 8-10 Å!

Cheers

Schara
Dr. Schara Safarian
Max Planck Institut für Biophysik 
Abteilung für Molekulare Membranbiologie

Max Planck Institute of Biophysics 
Department of Molecular Membrane Biology 

Max-von-Laue-Straße 3
60438 Frankfurt am Main
Deutschland/Germany

Twitter 
Google Scholar 



> Am 01.06.2020 um 07:18 schrieb Suchi Goel :
> 
> Hi Everyone,
> 
> I am new to this list. Have some problem of background in 2D classification, 
> that I can't get rid of for any parameter of regularization  (1.4-2.0) or 
> resolution limit (-1, no limit and 7-12 A limit). 
> Attached files are the output classes in Binning 2 and 1 respectively.
> Could some one help who has this experience. 
> 
> Best Regards,
> Suchi Goel
> 
> To unsubscribe from the CCP4BB list, click the following link:
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>  2020-06-01 at 14.07.37.png>




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Re: [ccp4bb] Completeness question

[Clemens Vonrhein]

Dear all,

On Sat, May 30, 2020 at 03:40:53PM +0100, Eleanor Dodson wrote:
> My pennysworth. If you find your maps look better after the
> anisotroy correction use it, but it may be helpful to those wo want to mine
> your data if you deposit the whole sphere..

Agree (which is what e.g. we provide when using STARANISO via autoPROC
[1]).

And in the same vein: those depositing isotropically truncated data
should consider also providing data to a higher diffraction limit to
give a potentially more accurate picture (if there is even a slight
indication of anisotropy - which there often is).

I find it very helpful even looking at an idealised (and therefore
simplified) picture of anisotropy as in

  http://staraniso.globalphasing.org/anisotropy_about.html

We can consider

 (1) for refinement:

 (1a) green+red, i.e. spherical (i.e. isotropically) truncated
  data

 (1b) green+blue, i.e. anisotropycally truncated data

 (2) for deposition:

 (2a) green+red => full sphere, but dropping real observations
  (blue)

 (2b) green+blue => all observations, but not providing
  insignificant/weak data (red) in all directions

 (2c) a sphere to the "tip" of blue (i.e. anisotropic diffraction
  limit) => all observations and all insignificant/weak data

Cheers

Clemens

[1] https://www.globalphasing.com/autoproc/ - which gives a mmCIF file
with (2a), (2b) and (2c) ready for deposition.


> eleanor
> 
> On Sat, 30 May 2020 at 09:36, Robbie Joosten 
> wrote:
> 
> > Hi Everyone,
> >
> > I've been looking at some recent PDB entries that have much lower
> > spherical) completeness than reported in the coordinate file. One reason
> > for this is that the data were anisotropicly truncated, another reason is
> > some mess-up with the deposition of the reflection data. There is a lot of
> > discussion about the former practice and I don't want to go in to that, but
> > the second one is obviously an error. Now how do I distinguish these cases?
> >
> > Sometimes, you can look at the reported number of reflections and compare
> > that to the deposited reflection file and you will find that something has
> > clearly gone wrong. However, the reported number of reflections is not
> > entirely reliable because of other issues so I'd rather not use it. If you
> > use PDBpeep (e.g. for 6rjy) you can see something is wrong, but that is
> > completely visual. Is there a tool in CCP4 that reports both spherical and
> > ellipsoidal completeness (on merged reflection data)? That would make it
> > easy to distinguish such cases.
> >
> > Cheers,
> > Robbie
> >
> > 
> >
> > To unsubscribe from the CCP4BB list, click the following link:
> > https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1
> >
> > This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a
> > mailing list hosted by www.jiscmail.ac.uk, terms & conditions are
> > available at https://www.jiscmail.ac.uk/policyandsecurity/
> >
> 
> 
> 
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> 
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-- 

*--
* Clemens Vonrhein, Ph.D. vonrhein AT GlobalPhasing DOT com
* Global Phasing Ltd., Sheraton House, Castle Park 
* Cambridge CB3 0AX, UK   www.globalphasing.com
*--



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Re: [ccp4bb] Completeness question

[vincent Chaptal]

Good evening,

since I've been contacted off line several times about this, I'm posting 
here the protocol to combine maps with 2 mtz files of original and 
truncated data.
I went through this procedure as I wanted to include the maps, and this 
was the tricky part of the procedure in my hands. This way, everyone can 
look at the same maps when critically assessing a structure (especially 
in the context of redo-ing and severe anisotropy).


In a new directory, put all 3 mtz files (original Intentisties, merged 
staraniso reflections, maps from buster)


/PATH_TO-DIR/BUSTER/Refine45/autoPROC/.

aP_deposition_prep -p combine -r maps.mtz -f staraniso_output.mtz -a 
Original_Intensities.mtz >aP_deposition_prep.log


It creates combine_all.cif

This file has everything but it has a “refln” naming issue.

To fix the “refln” issue:

cat combine_all.cif | sed -e "s/refln.F_sigma /refln.F_meas_sigma /g" -e 
"s%[ ]* #.*from dataset.*%%g" | awk 
'/^#/{if(s){print;next};c++;a[c]=$0;next}/^data/{print;if(s)next;s=1;for(i=1;i<=c;i++){print 
a[i]}next}{print}' > combine_all_refln_fixed.cif


 DONE.


Best
Vincent
ps: again, the credit of this script goes to the globalphasing team who 
kindly helped when I asked them for help with it.




Le 01/06/2020 à 11:47, Clemens Vonrhein a écrit :

Dear all,

On Sat, May 30, 2020 at 03:40:53PM +0100, Eleanor Dodson wrote:

My pennysworth. If you find your maps look better after the
anisotroy correction use it, but it may be helpful to those wo want to mine
your data if you deposit the whole sphere..

Agree (which is what e.g. we provide when using STARANISO via autoPROC
[1]).

And in the same vein: those depositing isotropically truncated data
should consider also providing data to a higher diffraction limit to
give a potentially more accurate picture (if there is even a slight
indication of anisotropy - which there often is).

I find it very helpful even looking at an idealised (and therefore
simplified) picture of anisotropy as in

   http://staraniso.globalphasing.org/anisotropy_about.html

We can consider

  (1) for refinement:

  (1a) green+red, i.e. spherical (i.e. isotropically) truncated
   data

  (1b) green+blue, i.e. anisotropycally truncated data

  (2) for deposition:

  (2a) green+red => full sphere, but dropping real observations
   (blue)

  (2b) green+blue => all observations, but not providing
   insignificant/weak data (red) in all directions

  (2c) a sphere to the "tip" of blue (i.e. anisotropic diffraction
   limit) => all observations and all insignificant/weak data

Cheers

Clemens

[1] https://www.globalphasing.com/autoproc/ - which gives a mmCIF file
 with (2a), (2b) and (2c) ready for deposition.



eleanor

On Sat, 30 May 2020 at 09:36, Robbie Joosten 
wrote:


Hi Everyone,

I've been looking at some recent PDB entries that have much lower
spherical) completeness than reported in the coordinate file. One reason
for this is that the data were anisotropicly truncated, another reason is
some mess-up with the deposition of the reflection data. There is a lot of
discussion about the former practice and I don't want to go in to that, but
the second one is obviously an error. Now how do I distinguish these cases?

Sometimes, you can look at the reported number of reflections and compare
that to the deposited reflection file and you will find that something has
clearly gone wrong. However, the reported number of reflections is not
entirely reliable because of other issues so I'd rather not use it. If you
use PDBpeep (e.g. for 6rjy) you can see something is wrong, but that is
completely visual. Is there a tool in CCP4 that reports both spherical and
ellipsoidal completeness (on merged reflection data)? That would make it
easy to distinguish such cases.

Cheers,
Robbie



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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

http://www.appicom.cnrs.fr

http://mmsb.cnrs.fr/en/





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[ccp4bb] Question about small molecule crystallography

[Jiyuan Ke]

Hi Everyone,

I want to crystallize a small organic molecule. I have very limited
experience in small molecule crystallography. I found that the Crystal
Screen HT from the Hampton research is good for both small molecule and
macromolecule crystallization. Plan to set up a sitting drop screen just
like setting up protein crystallization. I don’t know if this is the proper
way to do it. Is the MRC sitting drop 2-well plate (HR3-083) used for
protein crystallization good for small molecule crystallization? Are there
any special plates used for small molecule crystallization? Is room
temperature ok or not?

For data collection, can I use the beamline for protein crystals to collect
data on small molecule crystals? Larger oscillation angle, shorter
exposure, reduced beam intensity?

For structure determination, is SHELXL the preferred software for solving
small molecule structures?

If anyone has experience in small molecule crystallography, please help.
Thanks!

Best Regards,

-- 

*Jiyuan Ke, Ph.D.*


Research Investigator

H3 Biomedicine Inc.

300 Technology Square, Floor 5

Cambridge, MA 02139

Phone: 617-252-3923

Email: jiyuan...@h3biomedicine.com

Website: www.h3biomedicine.com

-- 
[This e-mail message may contain privileged, confidential and/or 
proprietary information of H3 Biomedicine. If you believe that it has been 
sent to you in error, please contact the sender immediately and delete the 
message including any attachments, without copying, using, or distributing 
any of the information contained therein. This e-mail message should not be 
interpreted to include a digital or electronic signature that can be used 
to authenticate an agreement, contract or other legal document, nor to 
reflect an intention to be bound to any legally-binding agreement or 
contract.]



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Re: [ccp4bb] Question about small molecule crystallography

[Artem Evdokimov]

Hi

A small organic molecule is typically crystallized from organic solvents
(or water, if soluble) by means of at least three main techniques:

1. slow evaporation of solvent leading to supersaturation and eventual
crystallization
2. supersaturation at higher temperature followed by gradual drop in
temperature causing crystallization
3. counter-diffusion of an incompatible solvent to drop solubility of the
substance and cause crystallization

Many times, just leaving an NMR tube with a tiny hole in the plastic cap
for a week or so will cause crystals to form.

Schnobviously, some substances will not crystallize easily - some form
oils, amorphous precipitates, etc. and others will form liquid hydrated
forms or just plain decompose. If you have any specific questions please
don't hesitate to contact me in person. I've spent half of my PhD
crystallizing weird small molecules for fun and profit.

As to how to solve structures of small molecules - any synchrotron is a
massive overkill. Just get in touch with a University X-ray lab, many of
which still have functional small molecule instruments. SHELX is the
software of choice - of course! (I still have the blue/white polka dot
SHELX cup, it's one of my more treasured curios).

Artem
- Cosmic Cats approve of this message


On Mon, Jun 1, 2020 at 6:01 PM Jiyuan Ke 
wrote:

> Hi Everyone,
>
> I want to crystallize a small organic molecule. I have very limited
> experience in small molecule crystallography. I found that the Crystal
> Screen HT from the Hampton research is good for both small molecule and
> macromolecule crystallization. Plan to set up a sitting drop screen just
> like setting up protein crystallization. I don’t know if this is the proper
> way to do it. Is the MRC sitting drop 2-well plate (HR3-083) used for
> protein crystallization good for small molecule crystallization? Are there
> any special plates used for small molecule crystallization? Is room
> temperature ok or not?
>
> For data collection, can I use the beamline for protein crystals to
> collect data on small molecule crystals? Larger oscillation angle, shorter
> exposure, reduced beam intensity?
>
> For structure determination, is SHELXL the preferred software for solving
> small molecule structures?
>
> If anyone has experience in small molecule crystallography, please help.
> Thanks!
>
> Best Regards,
>
> --
>
> *Jiyuan Ke, Ph.D.*
>
>
> Research Investigator
>
> H3 Biomedicine Inc.
>
> 300 Technology Square, Floor 5
>
> Cambridge, MA 02139
>
> Phone: 617-252-3923
>
> Email: jiyuan...@h3biomedicine.com
>
> Website: www.h3biomedicine.com
>
>
>
>
>
>
> [This e-mail message may contain privileged, confidential and/or
> proprietary information of H3 Biomedicine. If you believe that it has been
> sent to you in error, please contact the sender immediately and delete the
> message including any attachments, without copying, using, or distributing
> any of the information contained therein. This e-mail message should not be
> interpreted to include a digital or electronic signature that can be used
> to authenticate an agreement, contract or other legal document, nor to
> reflect an intention to be bound to any legally-binding agreement or
> contract.]
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1
>



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Re: [ccp4bb] Question about small molecule crystallography

[Peat, Tom (Manufacturing, Parkville)]

Hello Jiyuan,

One small point to note- as Artem says, small molecule crystals are often 
generated out of solvents and these same solvents often melt the standard 
protein crystallisation plates, so be careful what you put into a plastic plate.

As Artem mentioned, synchrotrons are generally overkill for small molecule 
structures (although there are exceptions). In this case, I would like to plug 
for the Australian Synchrotron which has a dedicated small molecule 
crystallographer and a beamline set up for small molecule crystallography (we 
do some protein crystallography there too!). So there is help available for 
those that do want to use synchrotrons for small molecule structures.

cheers, tom

Tom Peat
Proteins Group
Biomedical Program, CSIRO
343 Royal Parade
Parkville, VIC, 3052
+613 9662 7304
+614 57 539 419
tom.p...@csiro.au


From: CCP4 bulletin board  on behalf of Artem Evdokimov 

Sent: Tuesday, June 2, 2020 8:07 AM
To: CCP4BB@JISCMAIL.AC.UK 
Subject: Re: [ccp4bb] Question about small molecule crystallography

Hi

A small organic molecule is typically crystallized from organic solvents (or 
water, if soluble) by means of at least three main techniques:

1. slow evaporation of solvent leading to supersaturation and eventual 
crystallization
2. supersaturation at higher temperature followed by gradual drop in 
temperature causing crystallization
3. counter-diffusion of an incompatible solvent to drop solubility of the 
substance and cause crystallization

Many times, just leaving an NMR tube with a tiny hole in the plastic cap for a 
week or so will cause crystals to form.

Schnobviously, some substances will not crystallize easily - some form oils, 
amorphous precipitates, etc. and others will form liquid hydrated forms or just 
plain decompose. If you have any specific questions please don't hesitate to 
contact me in person. I've spent half of my PhD crystallizing weird small 
molecules for fun and profit.

As to how to solve structures of small molecules - any synchrotron is a massive 
overkill. Just get in touch with a University X-ray lab, many of which still 
have functional small molecule instruments. SHELX is the software of choice - 
of course! (I still have the blue/white polka dot SHELX cup, it's one of my 
more treasured curios).

Artem
- Cosmic Cats approve of this message


On Mon, Jun 1, 2020 at 6:01 PM Jiyuan Ke 
mailto:jiyuan...@h3biomedicine.com>> wrote:
Hi Everyone,

I want to crystallize a small organic molecule. I have very limited experience 
in small molecule crystallography. I found that the Crystal Screen HT from the 
Hampton research is good for both small molecule and macromolecule 
crystallization. Plan to set up a sitting drop screen just like setting up 
protein crystallization. I don’t know if this is the proper way to do it. Is 
the MRC sitting drop 2-well plate (HR3-083) used for protein crystallization 
good for small molecule crystallization? Are there any special plates used for 
small molecule crystallization? Is room temperature ok or not?

For data collection, can I use the beamline for protein crystals to collect 
data on small molecule crystals? Larger oscillation angle, shorter exposure, 
reduced beam intensity?

For structure determination, is SHELXL the preferred software for solving small 
molecule structures?

If anyone has experience in small molecule crystallography, please help.  
Thanks!

Best Regards,

--

Jiyuan Ke, Ph.D.


Research Investigator

H3 Biomedicine Inc.

300 Technology Square, Floor 5

Cambridge, MA 02139

Phone: 617-252-3923

Email: jiyuan...@h3biomedicine.com

Website: www.h3biomedicine.com

[https://docs.google.com/uc?export=download&id=0B1VBLPNVMUntenZVa3RFS3JOcVU&revid=0B1VBLPNVMUntbExRa0hyYWpCZGVNeFhZc0JwVit3bzU5c1Z3PQ]





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information of H3 Biomedicine. If you believe that it has been sent to you in 
error, please contact the sender immediately and delete the message including 
any attachments, without copying, using, or distributing any of the information 
contained therein. This e-mail message should not be interpreted to include a 
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contract or other legal document, nor to reflect an intention to be bound to 
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Thi