Re: [ccp4bb] XQuartz and High Sierra OSX

[Ruud Hovius]

Hello,

I had probelms this week with
1) Pymol 2.0 where clicking on the different  legacy plugins (except 
lighting) caused shut down of Pymol 2.0

2) CCP4 old style aborted upon re-start-up (which I do far too rarely...).

Upon restarting the mac,it appeared that several "Updates" of I do not 
know what got installed, and now CCP4 is fine, and Pymol too except the 
APBS electrostatics


If there is a solution for the electrostatics I would be happy to learn 
about this.


Greetings, Ruud


On 2/12/17 01:04, Steiner, Roberto wrote:

Hi all

Since my recent upgrade to High Sierra, XQuartz (most recent version 
2.7.11 - fresh installation) does not longer behave properly. 
Essentially, X11 does not seem to understand touchpad actions (for 
example cannot close X11 windows, cannot drag them around, etc.). This 
problems also affects programs that rely on X11. Has anyone 
experienced (and ideally solved) this problem?


Thanks in advance
Roberto

Roberto A. Steiner, PhD
Randall Centre of Cell and Molecular Biophysics
Faculty of Life Sciences and Medicine
King's College London

roberto.stei...@kcl.ac.uk 
Phone 0044 20 78488216
Fax    0044 20 78486435

Room 3.10A
New Hunt's House
Guy's Campus
SE1 1UL
London



--

Ruud Hovius
EPFL SB ISIC LIP
BCH 4209
CH-1015 Lausanne
+41-21-693-9442
http://people.epfl.ch/ruud.hovius

Re: [ccp4bb] new ContaMiner features

[Diana Tomchick]

It is also possible that it co-purifies via a nickel IMAC column purification 
step. The affinity of many E. coli proteins for such columns is well known.

Diana

**
Diana R. Tomchick
Professor
Departments of Biophysics and Biochemistry
University of Texas 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 Dec 1, 2017, at 6:31 PM, Ivan Shabalin  
wrote:

Dear All,

To add my voice to those who wrote crystallization artifacts happen - we just 
witnessed one today. A postdoc from another lab tried crystallizing a protein 
for months. Today, during data collection from his poorly diffracting crystals, 
one of our guys (Dr. Porebski) scaled the data and checked if there is a 
structure with the same Sp Gr and unit cell parameters in the PDB. And there 
was one - 4ZNZ (Escherichia coli carbonic anhydrase (YadF) in complex with Zn - 
an artifact of purification), deposited by our group two years ago. Quick 
molecular replacement showed that it is indeed the protein we had in the beam 
today.

The protein looked good on the SDS gel. The band for YadF was definitely less 
than 1% of the protein sample (less than one percent!!! if the gel would not be 
overloaded, nobody would see the band), but it still crystallized. We speculate 
that YadF from Escherichia coli was co-purified with the target protein due to 
relatively strong protein-protein interactions despite multiple purification 
steps.

We did not use ContaMiner, though. Instead, we just used the search of the unit 
cell parameters in the PDB, which is much faster (but works only if the 
structure of this particular artifact in the same SpGr is in PDB! otherwise, 
one should use ContaMiner or similar service). This feature is implemented in 
HKL3000, but it can also be done quickly on the PDB webpage. The procedure, as 
well as cases with other crystallization artifacts, is described in:

Protein purification and crystallization artifacts: The tale usually not told. 
(2016) Protein Sci. 25: 720-733

Today's example clearly shows that depositing these artifacts can greatly help 
others (it took just a few minutes to realize we had an artifact). Therefore, I 
would like to encourage everyone to deposit their artifacts to the PDB, 
especially if these artifacts were crystallized with unit cell parameters not 
reported previously. An increase in the size of the library of crystallization 
artifact structures deposited to the PDB can make troubleshooting of new 
artifacts much easier, and save much effort for those who are new (or not very 
new!) to such cases.


With best regards,
Ivan Shabalin, Ph.D.
Research Scientist,
Department of Molecular Physiology and Biological Physics,
University of Virginia,
1340 Jefferson Park Avenue, Pinn Hall,Room 4223,
Charlottesville, VA 22908

On 11/23/2017 02:35 PM, r...@mrc-lmb.cam.ac.uk wrote:
> Dear Stefan,
> Just a couple of thoughts:
> - first of all I think that Gerard is absolutely right, it would have been
> nice to raise such issues first with the developers. In my experience,
> Staraniso does a fantastic job if used correctly.
> - but if you're OK with public trials, may I ask: why on Earth would anybody
> need ContaMiner? Are you trying to offer some sort of computational cure for
> sloppy biochemistry? There is zero point in crystallizing crap samples, sorry
> to say this. In my 17 or so years in Strubi I've never heard of anybody
> crystallizing a "contaminant", being it a purification tag or whatever.
> I suppose this might have happened to somebody you know, hence the motivation
> to spend time on the bizarre ContaMiner. Which is a pity, a silly outcome
> would only teach people to do their job (or train their robots) properly.
> Best wishes,
> Radu






UT Southwestern


Medical Center



The future of medicine, today.

Re: [ccp4bb] new ContaMiner features

[Ivan Shabalin]

Dear All,

To add my voice to those who wrote crystallization artifacts happen - we 
just witnessed one today. A postdoc from another lab tried crystallizing 
a protein for months. Today, during data collection from his poorly 
diffracting crystals, one of our guys (Dr. Porebski) scaled the data and 
checked if there is a structure with the same Sp Gr and unit cell 
parameters in the PDB. And there was one - 4ZNZ (Escherichia coli 
carbonic anhydrase (YadF) in complex with Zn - an artifact of 
purification), deposited by our group two years ago. Quick molecular 
replacement showed that it is indeed the protein we had in the beam today.


The protein looked good on the SDS gel. The band for YadF was definitely 
less than 1% of the protein sample (less than one percent!!! if the gel 
would not be overloaded, nobody would see the band), but it still 
crystallized. We speculate that YadF from Escherichia coli was 
co-purified with the target protein due to relatively strong 
protein-protein interactions despite multiple purification steps.


We did not use ContaMiner, though. Instead, we just used the search of 
the unit cell parameters in the PDB, which is much faster (but works 
only if the structure of this particular artifact in the same SpGr is in 
PDB! otherwise, one should use ContaMiner or similar service). This 
feature is implemented in HKL3000, but it can also be done quickly on 
the PDB webpage. The procedure, as well as cases with other 
crystallization artifacts, is described in:


Protein purification and crystallization artifacts: The tale usually not 
told. (2016) Protein Sci. 25: 720-733


Today's example clearly shows that depositing these artifacts can 
greatly help others (it took just a few minutes to realize we had an 
artifact). Therefore, I would like to encourage everyone to deposit 
their artifacts to the PDB, especially if these artifacts were 
crystallized with unit cell parameters not reported previously. An 
increase in the size of the library of crystallization artifact 
structures deposited to the PDB can make troubleshooting of new 
artifacts much easier, and save much effort for those who are new (or 
not very new!) to such cases.



With best regards,
Ivan Shabalin, Ph.D.
Research Scientist,
Department of Molecular Physiology and Biological Physics,
University of Virginia,
1340 Jefferson Park Avenue, Pinn Hall,Room 4223,
Charlottesville, VA 22908

On 11/23/2017 02:35 PM, r...@mrc-lmb.cam.ac.uk wrote:

Dear Stefan,

Just a couple of thoughts:

- first of all I think that Gerard is absolutely right, it would have been
nice to raise such issues first with the developers. In my experience,
Staraniso does a fantastic job if used correctly.

- but if you're OK with public trials, may I ask: why on Earth would anybody
need ContaMiner? Are you trying to offer some sort of computational cure for
sloppy biochemistry? There is zero point in crystallizing crap samples, sorry
to say this. In my 17 or so years in Strubi I've never heard of anybody
crystallizing a "contaminant", being it a purification tag or whatever.

I suppose this might have happened to somebody you know, hence the motivation
to spend time on the bizarre ContaMiner. Which is a pity, a silly outcome
would only teach people to do their job (or train their robots) properly.

Best wishes,

Radu

[ccp4bb] XQuartz and High Sierra OSX

[Steiner, Roberto]

Hi all

Since my recent upgrade to High Sierra, XQuartz (most recent version 2.7.11 - 
fresh installation) does not longer behave properly. Essentially, X11 does not 
seem to understand touchpad actions (for example cannot close X11 windows, 
cannot drag them around, etc.). This problems also affects programs that rely 
on X11. Has anyone experienced (and ideally solved) this problem?

Thanks in advance
Roberto

Roberto A. Steiner, PhD
Randall Centre of Cell and Molecular Biophysics
Faculty of Life Sciences and Medicine
King's College London

roberto.stei...@kcl.ac.uk
Phone 0044 20 78488216
Fax0044 20 78486435

Room 3.10A
New Hunt's House
Guy's Campus
SE1 1UL
London

Re: [ccp4bb] Electrostatic Potential: Poisson-Boltzmann

[Robbie Joosten]

If you cannot trust the surface of your protein, perhaps you should not look at 
the the potential on the surface. Instead you can look at the field around your 
protein. This is less precise, but also less sensitive to local errors. If you 
want to know how your peptide finds your protein, this is actually more 
informative anyway.

There must be several programs that do this. I have done this for MHC in the 
past with YASARA. It really explained nicely how he peptide moved in.



Cheers,

Robbie



Sent from my Windows 10 phone




From: CCP4 bulletin board  on behalf of Dale Tronrud 

Sent: Friday, December 1, 2017 7:29:01 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Electrostatic Potential: Poisson-Boltzmann

   These are not easy questions to answer.  Certainly atoms,
particularly ones that are charged, even with fractional charges, have a
strong effect on the ESP.  If you delete them because you don't know
exactly where they are you will get a different answer than if you put
them in in some reasonable but unsupported location (as you have found).
 This result indicates that the peptide does affect the ESP
significantly and you have to consider it.

   You could build lots of models with the peptide in different
conformations and average all the maps.  This misses the point.  You
have uncertainty in your model which means that you have uncertainty in
your electrostatic potential.  Any particular ESP that you calculate and
draw conclusions from will have a large uncertainty and you must
consider that uncertainty when deciding between your potential
conclusions.  (I'm not sure if the pun is intended or not!)

   I suppose you could believe that each possible conformation exists to
some extent in reality which means that all the ESP's you calculate
exist in some fraction of the molecules in the cell.  It is possible
that only the molecules with a particular conformation of this peptide
have the ESP that allows the molecule to function.  Life is hard.

   Another issue that you must consider: If the exact conformation of
this loop causes changes to the ESP that you consider significant to
your understanding of the function of this protein, the presence and
conformation of neighboring proteins and solvent will also cause
significant changes to the ESP.  The biological context of the protein
becomes important.  If your interpretation depends critically on the
value and distribution of ESP then I'm not sure you can work this out
based on calculated ESP, considering the large uncertainty.

Dale Tronrud


On 12/1/2017 10:02 AM, chemocev marker wrote:
> Hi
>
> I am calculating the Electrostatic Potential of my protein. But there
> were few flexible region with high B-factor and I deleted that part of
> the protein and then recalculated it. But there I can see a big
> change.As I have a structure in the presence and the absence of the
> peptide and the these flexible regions have a better map in the
> structure without peptide and with peptide I have to delete them.
> I have a question, should I model these missing regions
>
> or
>
> I should ignor them
>
> best
>
> Jiri

Re: [ccp4bb] Do you have this type of diffraction pattern?

[James Phillips]

I agree with Roger Rowlett, try a room temp capillary mount.  Also ,what is
the magnification of the microscope/TV system you are using to center these
tiny crystals? Centering the loop is not enough.

James Phillips

On Fri, Dec 1, 2017 at 8:48 AM, Thomas, Leonard M.  wrote:

> Looking a bit closer are you sure the crystal is protein?  I see a couple
> of spots which could indicate a small molecule of some variety.
>
> Len
>
>
> On Nov 30, 2017, at 3:04 PM, Geny  wrote:
>
> ​
>
> Dear EveryOne,
> ​
> ​ I recently tried to collect the diffraction data, but unluckily, the
> diffraction patter is attached in this email. There is no any diffraction
> spots ​at all. It is my honor to ask experts worldwide why the crystal does
> this, and how to improve the quality of this kind of crystal. if you have
> any ideas, would you like to share?  Thank you so much and really
> appreciate it.
>
> Bests,
> Geny
>
>
> 
> 
>
>
> Leonard Thomas
> lmtho...@ou.edu
>
>
>
>

Re: [ccp4bb] Electrostatic Potential: Poisson-Boltzmann

[Dale Tronrud]

   These are not easy questions to answer.  Certainly atoms,
particularly ones that are charged, even with fractional charges, have a
strong effect on the ESP.  If you delete them because you don't know
exactly where they are you will get a different answer than if you put
them in in some reasonable but unsupported location (as you have found).
 This result indicates that the peptide does affect the ESP
significantly and you have to consider it.

   You could build lots of models with the peptide in different
conformations and average all the maps.  This misses the point.  You
have uncertainty in your model which means that you have uncertainty in
your electrostatic potential.  Any particular ESP that you calculate and
draw conclusions from will have a large uncertainty and you must
consider that uncertainty when deciding between your potential
conclusions.  (I'm not sure if the pun is intended or not!)

   I suppose you could believe that each possible conformation exists to
some extent in reality which means that all the ESP's you calculate
exist in some fraction of the molecules in the cell.  It is possible
that only the molecules with a particular conformation of this peptide
have the ESP that allows the molecule to function.  Life is hard.

   Another issue that you must consider: If the exact conformation of
this loop causes changes to the ESP that you consider significant to
your understanding of the function of this protein, the presence and
conformation of neighboring proteins and solvent will also cause
significant changes to the ESP.  The biological context of the protein
becomes important.  If your interpretation depends critically on the
value and distribution of ESP then I'm not sure you can work this out
based on calculated ESP, considering the large uncertainty.

Dale Tronrud


On 12/1/2017 10:02 AM, chemocev marker wrote:
> Hi
> 
> I am calculating the Electrostatic Potential of my protein. But there
> were few flexible region with high B-factor and I deleted that part of
> the protein and then recalculated it. But there I can see a big
> change.As I have a structure in the presence and the absence of the
> peptide and the these flexible regions have a better map in the
> structure without peptide and with peptide I have to delete them.
> I have a question, should I model these missing regions
> 
> or
> 
> I should ignor them
> 
> best
> 
> Jiri

[ccp4bb] Electrostatic Potential: Poisson-Boltzmann

[chemocev marker]

Hi

I am calculating the Electrostatic Potential of my protein. But there were
few flexible region with high B-factor and I deleted that part of the
protein and then recalculated it. But there I can see a big change.As I
have a structure in the presence and the absence of the peptide and the
these flexible regions have a better map in the structure without peptide
and with peptide I have to delete them.
I have a question, should I model these missing regions

or

I should ignor them

best

Jiri

[ccp4bb] -Postdoc Position in Structural Biology at Baylor College of Medicine, Houston, Texas

[Qin, Liying]

Dear all,

Hello! I am posting a job position on behalf of Dr. Choel Kim. Below is the 
description for the job. Should you have any question regarding the position, 
please directly contact Dr. Kim at c...@bcm.edu.


An NIH funded postdoctoral research scientist position is available at Dr. 
Choel Kim’s laboratory in the Department of Pharmacology and Chemical Biology 
at Baylor College of Medicine.  The successful candidate will participate in 
our structure based drug discovery program targeting signaling proteins in the 
Nitric Oxide-cGMP signaling cascade using X-ray crystallography and Cryo-EM. 
These enzymes modulate the cellular level of cGMP or the responses to this 
crucial secondary messenger representing therapeutic targets for treating many 
hypertensive diseases, such as arterial and pulmonary hypertension, heart 
failure and erectile dysfunction.

The successful candidates must have PhD degree and have strong skills in 
molecular biology, protein expression/purification and macromolecular 
crystallography. Preference will be given to those who have a strong drive and 
motivation. The position includes ample opportunities for training in Cryo-EM. 
Experience with Cryo-EM is not necessary, but beneficial.

The Department of Pharmacology and Chemical Biology provides state of the art 
facilities for Biophysics and X-ray crystallography and Cryo-EM. Salary and 
benefits are offered based on NIH guidelines.  Applicants should submit 
curriculum vitae, cover letter outlining career goals and contact information 
for 3 references to Dr. Choel Kim (c...@bcm.edu).



Best regards,
Liying



Liying Qin
Ph.D. Candidate
BMB Program
Baylor College of Medicine
One Baylor Plaza,Houston, TX77030, USA

Re: [ccp4bb] Do you have this type of diffraction pattern?

[Thomas, Leonard M.]

Looking a bit closer are you sure the crystal is protein?  I see a couple of 
spots which could indicate a small molecule of some variety.

Len


On Nov 30, 2017, at 3:04 PM, Geny > 
wrote:

​

Dear EveryOne,
​
​ I recently tried to collect the diffraction data, but unluckily, the 
diffraction patter is attached in this email. There is no any diffraction spots 
​at all. It is my honor to ask experts worldwide why the crystal does this, and 
how to improve the quality of this kind of crystal. if you have any ideas, 
would you like to share?  Thank you so much and really appreciate it.

Bests,
Geny





Leonard Thomas
lmtho...@ou.edu

[ccp4bb] PhD positions-Structural Biology, Barcelona

[Esther Fernandez]

 *PhD positions-Structural Biology*

We are pleased to inform you that the “*IRB Barcelona International PhD
Fellowships Programme 2018”* call is now open.

For the 2018-2019 academic year, *IRB Barcelona is offering 10 PhD
fellowships, including Structural Biology*,  for young scientists from the
national and international community.
We encourage applications from highly motivated graduates with outstanding
qualifications in *biology, biochemistry, physics, pharmacology, chemistry
or related fields*. Successful candidates will join research groups headed
by top-level scientists and will carry out their research in cutting-edge
fields of biomedicine in a stimulating environment.

The deadline for applications is *7 January 2018, 15:00h CET Time.*

For further information, please visit our website

or
alternatively contact us at p...@irbbarcelona.org