Re: [ccp4bb] ligand refinemnet

[Kontopidis George]

Peter from you description seems that big blob is not your ligand. In 3.0
resolution molecules with similar size, including ligands, tend to look
similar in shape as well. I suggest you refine further your structure and
check again later this N-terminal blob of density. It may tern out to be a
bias from your molecular replacement model. If it is still there I will
suggest to rum a MS and compare the MW with the one from your protein
structure. That will give you an indication that probable you have something
attached to N-terminal of your protein.

George 

 

 

  _  

From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of peter
hudson
Sent: Thursday, April 02, 2009 9:55 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] ligand refinemnet

 

Hello all

I have refined and built model a dataset of 3.0A resolution dataset. This
model is assocaied with a ligand. After final refinement and model building
i found  a big blob of  Fo-Fc density of around 4sigma level at the
N-terminal of the fianl model. My protein doesnot carry any tag at the
N-terminal. But, the template i have used for the molecular replacement
carrying a tag at the N-termianl, which can occupy only 1/4th of this
positive density after superpostion. My crystallisation conditions component
cannot occupy such higher level of positive density. Since my protein binds
to a ligand, i looked carefully to the positive density and it seems very
similar to the ligand density but its obscure. Refinement after fitting the
ligand leads to very high B-factor of  the ligan, which vary for various
atoms from 50-90 and simultaneously positive density goes off. I also tried
to change the occupancy of the ligand but as i reduce the occupancy the B
factor comes down at the noraml expected average B factor value but again
Fo-Fc density appears in the map over ligand. If i leave to refine the
occupancy to the programme automatically, this lead to the zero occupancy of
the few atoms in the ligand and avearge B factor remains normal.
suggestions  would be appreciated.

Thanks in advance
Peter 

Re: [ccp4bb] ligand refinemnet

[Tim Gruene]

Hello Peter,

at 3A resolution occupancy refinement is supposed to fail, I'd say.
The large B-values can make up for reduced occupancy (as you already 
pointed out to) but also large flexibility.


You might try ten refinement runs in parallel with fixed occupancies 
varying from 10% to 100% and check whether you can detect signs of a local 
minimum at a certain occupancy - e.g. real space CC, best fit of B-values, 
etc. Yet much more you can probably not do.


If you cannot come to a satisfactory solution you may place a model ligand 
and describe the situation in the PDB header. That's not ideal but 
probably one of the best ways to go in my opinion.


Tim

--
Tim Gruene
Institut fuer anorganische Chemie
Tammannstr. 4
D-37077 Goettingen

GPG Key ID = A46BEE1A


On Thu, 2 Apr 2009, peter hudson wrote:


Hello all

I have refined and built model a dataset of 3.0A resolution dataset. This
model is assocaied with a ligand. After final refinement and model
building i found  a big blob of  Fo-Fc density of around 4sigma level at
the N-terminal of the fianl model. My protein doesnot carry any tag at
the N-terminal. But, the template i have used for the molecular
replacement carrying a tag at the N-termianl, which can occupy only 1/4th
of this positive density after superpostion. My crystallisation
conditions component cannot occupy such higher level of positive density.
Since my protein binds to a ligand, i looked carefully to the positive
density and it seems very similar to the ligand density but its obscure.
Refinement after fitting the ligand leads to very high B-factor of  the
ligan, which vary for various atoms from 50-90 and simultaneously
positive density goes off. I also tried to change the occupancy of the
ligand but as i reduce the occupancy the B factor comes down at the
noraml expected average B factor value but again Fo-Fc density appears in
the map over ligand. If i leave to refine the occupancy to the programme
automatically, this lead to the zero occupancy of the few atoms in the
ligand and avearge B factor remains normal.  suggestions  would be
appreciated.

Thanks in advance
Peter

Re: [ccp4bb] Lowest resolution you can do MR with

[Bjørn Panyella Pedersen]

I guess it depends on your criteria for success. We made a successful MR using 
data to 8Å with a search-model with 20% identity covering 90% of the target. The 
resultant phases gave a map where a few new 'blobs' could be observed. Not very 
useful, but the MR-phases could be used to solve the HA substructure of a 
derivative dataset we had using anomalous Fourier analysis. Thus the low-res MR 
solution lead to the success of our _experimental_ phasing.
Especially pay attention on the quality of your low-res data. In our hands the 
best dataset for MR was not the best dataset for later building the structure. 
Test different programs (Phaser worked for us, but might struggle if you have 
translational ncs (I think?)). Remember to play with the input parameters 
(especially resolution cutoff, search-model, rms of search). Our MR first 
succeeded after ~100 runs using different parameters (scripting is useful here!).


Good luck :)
-Bjørn


Muthiah wrote:
What is the lowest resolution one can try to do molecular replacement 
with? I have a 3.6 angstroms resolution data for a protein-DNA complex 
and wondering whether I can try MR to see the density for DNA.


--
Bjørn Panyella Pedersen, PhD
Postdoc

PUMPKIN, Centre for Membrane Pumps in Cells and Disease
University of Aarhus, Dept. Molecular Biology
Gustav Wieds Vej 10C DK-8000 Aarhus C, Denmark

Phone: +45 89425261
E-mail: bj...@bioxray.au.dk
http://www.bioxray.au.dk

[ccp4bb] Vacancies at Diamond Light Source

[Hannon, TJ (Tim)]

Dear CCP4BB Members,
 
Diamond have a number of vacancies, please click on the links below for
details.
 
Best regards,
 
Tim Hannon
Diamond Human Resources
01235 778452
 
Principal Beamline Scientists
http://www.diamond.ac.uk/Jobs/CurrentVacancies/Scientific/DIA0488.htm
 
http://www.diamond.ac.uk/Jobs/CurrentVacancies/Scientific/DIA0489.htm
 
http://www.diamond.ac.uk/Jobs/CurrentVacancies/Scientific/DIA0490.htm
 
Group Leader
http://www.diamond.ac.uk/Jobs/CurrentVacancies/Scientific/DIA0506SB.htm
 
Beamline Scientist
http://www.diamond.ac.uk/Jobs/CurrentVacancies/Scientific/DIA0495.htm
 
PDRA
http://www.diamond.ac.uk/Jobs/CurrentVacancies/Scientific/DIA0507.htm
 
All Scientific Vacancies
http://www.diamond.ac.uk/Jobs/CurrentVacancies/Scientific/default.htm

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are not the intended addressee or an authorised recipient of the addressee 
please notify us of receipt by returning the e-mail and do not use, copy, 
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Any opinions expressed within this e-mail are those of the individual and not 
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Diamond Light Source Limited (company no. 4375679). Registered in England and 
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Innovation Campus, Didcot, Oxfordshire, OX11 0DE, United Kingdom

 

[ccp4bb]

[Marc Graille]

Hi,
We had success using a testosterone hemisuccinate derivative which is 
much more soluble.
For details, see: Drevelle et al; J Mol Biol. 'jour', 'J Mol Biol.');> 2006 Apr 28;358(2):455-71


Hope this helps!!
Marc

KUMARASWAMI MUTHIAH a écrit :
Anybody tried to cocrystallize the protein-progesterone or estrogen 
complexes, if so how do you go about the solubility of these 
compounds? Progesterone is only soluble in 50% chloroform or 100% DMSO 
and the dilution of this stock is not possible as chloroform falls out 
of solution. Lots of papers out there used soaking with progesterone 
or expressed the protein in the presence of progesterone. Any 
suggestions would be appreciated.

Thanks




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--
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Yeast Structural Genomics, IBBMC CNRS UMR8619 Bat 430 Universite Paris Sud 
91405 Orsay Cedex
NEW PHONE NUMBER
Tel: (33) 1 69 15 31 57
Fax: (33) 1 69 85 37 15

[ccp4bb] CCP4 tcl installation problem (mac)

[Simon Kolstoe]

Dear ccp4bb,

I'm trying to update Tcl/Tk on a mac running OSX5.6 as my version of  
mosflm has suddenly started crashing when trying to autoindex, so I  
figured the best thing to do was to reinstall both mosflm and its  
dependencies.


So I went to the CCP4 download page and used the Daresbury ftp site to  
get the file "Tcl-Tk++-osx-universal.dmg.gz" which I then unzip, mount  
and run without a problem. HOWEVER when I open a new terminal and type  
"wish" my previous version (Tcl 8.4.10) opens instead of the new  
version, 8.4.18. I'm guessing that I need to point the soft-link in  
my /usr/local/bin directory to the new version, but I can't find where  
it has installed on my machine! If anyone could give me a hand and  
tell me where CCP4's Tcl installs itself I  would be grateful.


Thanks,

Simon

Re: [ccp4bb] Eleven plausible phasing elements remain unused

[Thomas Womack]

On Wed, 2009-04-01 at 14:33 -0700, Ethan Merritt wrote:
> On Wednesday 01 April 2009 07:21:16 Thomas Womack wrote:
> > A perusal of the PDB reveals that the game of Periodic Table bingo still
> > has eleven rounds to run:
> > 
> > scandium, titanium, germanium, zirconium, niobium, neodymium,
> > dysprosium, thulium, hafnium, bismuth and thorium remain absent from PDB
> > entries.
> 
> Does this imply that there is a PDB entry containing Radon?

I defined 'plausible' as a half-life greater than a billion years,
though I wouldn't have been totally amazed to see a plutonium or
technetium derivative.  Elements with half-lives 10^6 to 10^9 years for
the most stable isotope are Np, Tc, Cm, Pu; next shortest is 31kyears
for 231Pa.  The long-lived curium-247 and plutonium-244 isotopes are
neutron-heavy and inconvenient to produce.

The web-accessible subset of the ICSD features a technetium arsenide, a
plutonium boride, a sodium neptunate(VII) and an americium iodide.

Tom

Re: [ccp4bb] RIP SGI

[Thomas, Leonard M.]

We were just talking about SGI yesterday while getting rid of the last of old 
Indigo 2's.  Somebody asked if they were still in business and I said I think 
so. I guess I will have to change my answer not.  They were great at the time 
but like a lot of companies in the trash heap they tried to live on there past 
and did not really look ahead, see Digital.


Leonard M. Thomas Ph.D.
University of Oklahoma
Department of Chemistry and Biochemistry
620 Parrington Oval
Norman, OK 73072
405-325-1126
lmtho...@ou.edu

From: CCP4 bulletin board [ccp...@jiscmail.ac.uk] On Behalf Of William G. Scott 
[wgsc...@chemistry.ucsc.edu]
Sent: Wednesday, April 01, 2009 9:37 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] RIP SGI

Sadly, it is no joke:

Silicon Graphics timeline - San Jose Mercury News:

http://www.mercurynews.com/businessheadlines/ci_12049125

[ccp4bb] friction

[mesters]

Let me start by quoting the following..

"/Therefore I'm afraid that we (i.e. PDB) are under considerable 
pressure from the community at large to implement our publication policy 
in this area.

We do understand your concern to make the entry as accurate as possible."/

I love the /"under considerable pressure"/ and /"our publication policy"/?
I am for sure not exerting pressure and do have my problems with the 
policies at times.


Are you the one exerting pressure or do you have problems at times with 
the policies? Please let me know!


- J. -

p.s. In addition, over the years, the deposition in general has not 
become any simpler or faster. Or, has it?

--

Dr. Jeroen R. Mesters
Gruppenleiter Strukturelle Neurobiologie und Kristallogenese
Institut für Biochemie, Universität zu Lübeck
Zentrum für Medizinische Struktur- und Zellbiologie
Ratzeburger Allee 160, D-23538 Lübeck
Tel: +49-451-5004065, Fax: +49-451-5004068
Http://www.biochem.uni-luebeck.de
Http://www.iobcr.org
Http://www.selfish-brain.org
Http://www.opticryst.org
--
If you can look into the seeds of time and say
which grain will grow and which will not - speak then to me  (Macbeth)
--

[ccp4bb] Angle between two different helices

[Kim Henrick]

the program
  www.ebi.ac.uk/~henrick/doss.tar.gz
based on promotif  will do the angle between helices
(so then will promotif)

doss is f90 so may work with gfortran




--
Kim HENRICKhenr...@ebi.ac.uk ::telephone:  +44 (0) 1223 494629

Re: [ccp4bb] friction

[Anastassis Perrakis]

On Apr 2, 2009, at 16:04, mesters wrote:


Let me start by quoting the following..

"/Therefore I'm afraid that we (i.e. PDB) are under considerable
pressure from the community at large to implement our publication  
policy

in this area.
We do understand your concern to make the entry as accurate as  
possible."/


I love the /"under considerable pressure"/ and /"our publication  
policy"/?

I am for sure not exerting pressure and do have my problems with the
policies at times.

Are you the one exerting pressure or do you have problems at times  
with

the policies? Please let me know!



Since that is a bit out of context I do not see how I could comment ...


- J. -

p.s. In addition, over the years, the deposition in general has not
become any simpler or faster. Or, has it?


Thats a clear question ... Yes, in my opinion PDB deposition has  
changed over the years and is much better now than ever before.
Most of the people that have the rather dull job of checking entries  
are doing an excellent job and are friendly and understanding.
(I have dealt mostly with the EBI/MSD site for depositions so I cant  
tell for the RCSB)


A.

Re: [ccp4bb] GLRF and interpreting self rotation functions...

[Jeff Speir]

A standard orientation is anything you want it to be and is usually  
defined in the context of orthogonal axes.  It is simply a reference  
point from which you apply the results of your search.  We usually  
use one or more of the orthogonal axes as a starting point for easier  
visualization.


To define a 3-fold axis for a locked search in GLRF (if I understood  
your message correctly):


locsym 90.0 90.0 120.0 3 polar

This defines a standard orientation for that 3-fold.  The search will  
then move the 3-fold by the given parameters to new orientations and  
calculate the peak height at those grid points after applying 3-fold  
NCS.  Which axis this actually defines as your standard orientation  
and how it is then moved during the search depends on your choice of  
conventions for orthogonalization and polar angle searches.


Cheers,

Jeff


On Apr 1, 2009, at 3:32 PM, Francis E Reyes wrote:

I am experimenting with GLRF and am having trouble calculating the  
locked self  rotation function for a protein of known structure.  
The protein has a 3 fold NCS axis that is not parallel to a  
crystallographic axis. I'm at the step of specifying the local  
symmetry elements for the locked self rotation function.


I guess the goal here is to search for 'one general rotation which  
will bring the non crystallographic symmetry point group in a  
"standard" orientation. What does this mean? How do I write the  
LOCSymmetry instruction for the input file?


My s.g. is P 21 21 21 and the following solutions are found for the  
normal self rotation function.




Fine searches around peaks with the slow rotation function --

  The large-term cut-off is  1.50

 Listing of the fitted angles of the top5 peaks --

  No. Old Angles S A N G L  
E   O A N G L EOld Ht.New Ht.
 (polar,  
XYK)  (polar, XZK)
 phi  psi   
kap phi  psi  kap


1  52.000   50.000  120.000  52.000   50.500   
120.000  53.246  127.449  120.000417.36420.74
2 128.000  130.000  120.000 128.000  129.500   
120.000 233.246  127.449  120.000417.36420.74
3 128.000   50.000  120.000 129.000   50.000   
120.000 126.870  126.536  120.000400.32407.69
4  52.000  130.000  120.000  51.000  130.000   
120.000 306.870  126.536  120.000400.32407.68
5  44.000   60.000  120.000  42.000   61.000   
120.000  36.719  125.820  120.000385.56391.96


The goal of all this is to use GLRF to explore NCS in cases where  
structures are not known and molecular replacement


Thanks

FR

-
Francis Reyes M.Sc.
215 UCB
University of Colorado at Boulder

gpg --keyserver pgp.mit.edu --recv-keys 67BA8D5D

8AE2 F2F4 90F7 9640 28BC  686F 78FD 6669 67BA 8D5D

Re: [ccp4bb] friction or releasing pdb's

[Engin Ozkan]

Moving the subject further away from the original post...

I agree that the pdb deposition process has gotten better, but I still 
regularly have issues with releasing of newly published structures. 
There seem to be delays; just as you are reading this brand new, 
interesting structure, you realize that the pdb and structure factors 
are not released. Quite annoying. This is in large part the fault of the 
authors, but maybe PDB could do better, too.


Here is a funny case (in the spirit of Fools' Day). About a year ago, I 
asked the PDB to release a structure from a paper published in Nature. 
Their response was, they could not, because the publication was a letter 
to Nature, not an article. I rest my case.


Engin

On 4/2/09 7:56 AM, Anastassis Perrakis wrote:

On Apr 2, 2009, at 16:04, mesters wrote:


Let me start by quoting the following..

"/Therefore I'm afraid that we (i.e. PDB) are under considerable
pressure from the community at large to implement our publication policy
in this area.
We do understand your concern to make the entry as accurate as 
possible."/


I love the /"under considerable pressure"/ and /"our publication 
policy"/?

I am for sure not exerting pressure and do have my problems with the
policies at times.

Are you the one exerting pressure or do you have problems at times with
the policies? Please let me know!



Since that is a bit out of context I do not see how I could comment ...


- J. -

p.s. In addition, over the years, the deposition in general has not
become any simpler or faster. Or, has it?


Thats a clear question ... Yes, in my opinion PDB deposition has 
changed over the years and is much better now than ever before.
Most of the people that have the rather dull job of checking entries 
are doing an excellent job and are friendly and understanding.
(I have dealt mostly with the EBI/MSD site for depositions so I cant 
tell for the RCSB)


A.

Re: [ccp4bb] friction or releasing pdb's

[Phil Jeffrey]

PDB seem to take about a week to release coordinates that are HPUB (hold 
for publication) from when we ask them to.  Sometimes they drop the 
ball, but mostly this is what we see.  If I ask them to release the 
coordinates early to get around this lag, I can largely guarantee that 
I'll forget to update the JRNL reference.


I believe that the lead time is unnecessarily long.  Perhaps one day 
might be more reasonable.


Phil Jeffrey

Engin Ozkan wrote:
I agree that the pdb deposition process has gotten better, but I still 
regularly have issues with releasing of newly published structures. 
There seem to be delays; just as you are reading this brand new, 
interesting structure, you realize that the pdb and structure factors 
are not released. Quite annoying. This is in large part the fault of the 
authors, but maybe PDB could do better, too.

Re: [ccp4bb] friction or releasing pdb's

[Engin Ozkan]

I do not mind PDB releasing only on Wednesdays at a certain time of the 
day. What should happen is the authors/publishers should let PDB know 
the publication date before it becomes available (supposedly days or 
weeks before the actual publication, online or in print), and PDB can 
automatically release pdb/sf the week of the publication or the 
Wednesday before.


With requesting pdb releases, I take two routes: ask the journal and ask 
RCSB.  Asking journals used to work, although my last attempt was 
unsuccessful. If the journal agrees to do anything, they may let the 
authors or PDB know, so after a while, the release happens. Asking RCSB 
works if they can find the reference themselves, except in the bizarre 
case I cited.


But still, the reader should not have to search for data supposed to 
publish simultaneously with the article, right?


Engin

On 4/2/09 8:53 AM, Phil Jeffrey wrote:
PDB seem to take about a week to release coordinates that are HPUB 
(hold for publication) from when we ask them to.  Sometimes they drop 
the ball, but mostly this is what we see.  If I ask them to release 
the coordinates early to get around this lag, I can largely guarantee 
that I'll forget to update the JRNL reference.


I believe that the lead time is unnecessarily long.  Perhaps one day 
might be more reasonable.


Phil Jeffrey

Engin Ozkan wrote:
I agree that the pdb deposition process has gotten better, but I 
still regularly have issues with releasing of newly published 
structures. There seem to be delays; just as you are reading this 
brand new, interesting structure, you realize that the pdb and 
structure factors are not released. Quite annoying. This is in large 
part the fault of the authors, but maybe PDB could do better, too.

[ccp4bb] Identifying an unknown ligand

[Abhinav Kumar]

Hi,

I am refining a structure and have a region of unmodeled density into 
which I am trying to fit a ligand. The identity of the ligand is not 
obvious, so I modeled a bunch of dummy atoms into the density.
Could you please have a look at the map and pdb files and help me 
identify this ligand?


Please see http://smb.slac.stanford.edu/~abhinavk/UNL/unl.html

Thanks 
Abhinav 

Stanford Synchrotron Radiation Laboratory 
Joint Center for Structural Genomics 
Mail Stop 99 
Phone: (650) 926-2992 
Fax: (650) 926-3292 

Re: [ccp4bb] Identifying an unknown ligand

[Roger Rowlett]

Some context would be helpful (essential?). What's
in the crystallization solution? What, if anything, is known about the
protein of interest? What is the ligand interacting with (metal ions,
hydrogen bonding donors/acceptors, charged residues, etc?), and what
are the interaction distances? Do any of the atoms appear to be "heavy"
or show anomalous scattering, if applicable, at the wavelength used? 
etc. etc.

-- 

Roger S. Rowlett
Professor
Colgate University Presidential Scholar
Department of Chemistry
Colgate University
13 Oak Drive
Hamilton, NY 13346

tel: (315)-228-7245
ofc: (315)-228-7395
fax: (315)-228-7935
email: rrowl...@mail.colgate.edu

Abhinav Kumar wrote:

  Hi,

I am refining a structure and have a region of unmodeled density into 
which I am trying to fit a ligand. The identity of the ligand is not 
obvious, so I modeled a bunch of dummy atoms into the density.
Could you please have a look at the map and pdb files and help me 
identify this ligand?

Please see http://smb.slac.stanford.edu/~abhinavk/UNL/unl.html

Thanks 
Abhinav 

Stanford Synchrotron Radiation Laboratory 
Joint Center for Structural Genomics 
Mail Stop 99 
Phone: (650) 926-2992 
Fax: (650) 926-3292 
  

[ccp4bb] cryoprotectant for 1,6 hexanediol

[HanJie_Heng Chiat Tai]

Hi,

 

I have a crystal grown in 2.1M 1,6 hexanediol/0.1 M tro-sodium citrate (pH 6.5).

 

What's the cryoprotectant can be used to flash cool this crystal? 

 

Any online protein crystal cryoprotectant database or published literature 
available I can check with to determine to type and concentration of the 
cryoprotectant used for my crystal.


HengChiat Tai 



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Re: [ccp4bb] Identifying an unknown ligand

[Andrew Gulick]

My money is on:

Dithiothreitol or dithioerythritol.

Any chance they are in there? If not and you believe this copurified with
your protein, than I'd guess erythritol or threitol.

(I didn't bother trying to gauge the stereochemistry from your picture)

Cheers
Andy



On 4/2/09 1:38 PM, "Abhinav Kumar"  wrote:

> Hi,
> 
> I am refining a structure and have a region of unmodeled density into
> which I am trying to fit a ligand. The identity of the ligand is not
> obvious, so I modeled a bunch of dummy atoms into the density.
> Could you please have a look at the map and pdb files and help me
> identify this ligand?
> 
> Please see http://smb.slac.stanford.edu/~abhinavk/UNL/unl.html
> 
> Thanks 
> Abhinav 
> 
> Stanford Synchrotron Radiation Laboratory
> Joint Center for Structural Genomics
> Mail Stop 99 
> Phone: (650) 926-2992
> Fax: (650) 926-3292

-- 
Andrew M. Gulick, Ph.D.
---
(716) 898-8619
Hauptman-Woodward Institute
700 Ellicott St
Buffalo, NY 14203
---
Hauptman-Woodward Institute
Dept. of Structural Biology, SUNY at Buffalo

http://www.hwi.buffalo.edu/Faculty/Gulick/Gulick.html
http://labs.hwi.buffalo.edu/gulick

Re: [ccp4bb] Identifying an unknown ligand

[Abhinav Kumar]

Some more info about this structure:
Crystallization conditions: Glycerol, 0.1700M NaOAc, 25.5000% PEG-4000, 
0.1M TRIS pH 8.5


This does not sit on any crystalllographic symmetry axis.
But it sits right between two monomers and the NCS 2-fold axis.

The protein is chemotaxis protein CheX.

Environment: Some main chain amide Ns around it, as well as phobic side 
chains.


No peak in anomalous difference fourier map.


Thanks 
Abhinav 

Stanford Synchrotron Radiation Laboratory 
Joint Center for Structural Genomics 
Mail Stop 99 
Phone: (650) 926-2992 
Fax: (650) 926-3292 




Abhinav Kumar wrote:

Hi,

I am refining a structure and have a region of unmodeled density into 
which I am trying to fit a ligand. The identity of the ligand is not 
obvious, so I modeled a bunch of dummy atoms into the density.
Could you please have a look at the map and pdb files and help me 
identify this ligand?


Please see http://smb.slac.stanford.edu/~abhinavk/UNL/unl.html

Thanks Abhinav
Stanford Synchrotron Radiation Laboratory Joint Center for Structural 
Genomics Mail Stop 99 Phone: (650) 926-2992 Fax: (650) 926-3292 

Re: [ccp4bb] cryoprotectant for 1,6 hexanediol

[Jim Pflugrath]

The cryoprotectant is 1,6 hexanediol.

Jim

On Thu, 2 Apr 2009, HanJie_Heng Chiat Tai wrote:




Hi,



I have a crystal grown in 2.1M 1,6 hexanediol/0.1 M tro-sodium citrate (pH 6.5).



What's the cryoprotectant can be used to flash cool this crystal?



Any online protein crystal cryoprotectant database or published literature 
available I can check with to determine to type and concentration of the 
cryoprotectant used for my crystal.


HengChiat Tai



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Re: [ccp4bb] cryoprotectant for 1,6 hexanediol

[HanJie_Heng Chiat Tai]

Hi, Jim,

 

What's the concentration? I know that [hexanediol] between 2.5 - 3.4 M no 
additional cryoprotectant is required.

 

But in my case my hexanediol conc is only 2.1 M

Rgds,



HengChiat Tai (HanJie)


 

> Date: Thu, 2 Apr 2009 13:11:27 -0500
> From: jim.pflugr...@rigaku.com
> To: chemtai2...@hotmail.com
> CC: CCP4BB@JISCMAIL.AC.UK
> Subject: Re: [ccp4bb] cryoprotectant for 1,6 hexanediol
> 
> The cryoprotectant is 1,6 hexanediol.
> 
> Jim
> 
> On Thu, 2 Apr 2009, HanJie_Heng Chiat Tai wrote:
> 
> >
> >
> > Hi,
> >
> >
> >
> > I have a crystal grown in 2.1M 1,6 hexanediol/0.1 M tro-sodium citrate (pH 
> > 6.5).
> >
> >
> >
> > What's the cryoprotectant can be used to flash cool this crystal?
> >
> >
> >
> > Any online protein crystal cryoprotectant database or published literature 
> > available I can check with to determine to type and concentration of the 
> > cryoprotectant used for my crystal.
> >
> >
> > HengChiat Tai
> >
> >
> >
> > _
> > Rediscover Hotmail®: Get e-mail storage that grows with you.
> > http://windowslive.com/RediscoverHotmail?ocid=TXT_TAGLM_WL_HM_Rediscover_Storage1_042009

_
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Re: [ccp4bb] cryoprotectant for 1,6 hexanediol

[Jürgen Bosch]

If you are uncertain, just freeze your buffer :-)
Very old trick from the stone age of crystallography.

Long time ago ~ 1999 I cryoed one crystal with 1.5 M hexanediol but  
there was also 10% glycerol around.


Jürgen

On 2 Apr 2009, at 14:56, HanJie_Heng Chiat Tai wrote:


Hi, Jim,

What's the concentration? I know that [hexanediol] between 2.5 - 3.4  
M no additional cryoprotectant is required.


But in my case my hexanediol conc is only 2.1 M

Rgds,
HengChiat Tai (HanJie)




> Date: Thu, 2 Apr 2009 13:11:27 -0500
> From: jim.pflugr...@rigaku.com
> To: chemtai2...@hotmail.com
> CC: CCP4BB@JISCMAIL.AC.UK
> Subject: Re: [ccp4bb] cryoprotectant for 1,6 hexanediol
>
> The cryoprotectant is 1,6 hexanediol.
>
> Jim
>
> On Thu, 2 Apr 2009, HanJie_Heng Chiat Tai wrote:
>
> >
> >
> > Hi,
> >
> >
> >
> > I have a crystal grown in 2.1M 1,6 hexanediol/0.1 M tro-sodium  
citrate (pH 6.5).

> >
> >
> >
> > What's the cryoprotectant can be used to flash cool this crystal?
> >
> >
> >
> > Any online protein crystal cryoprotectant database or published  
literature available I can check with to determine to type and  
concentration of the cryoprotectant used for my crystal.

> >
> >
> > HengChiat Tai
> >
> >
> >
> > _
> > Rediscover Hotmail®: Get e-mail storage that grows with you.
> > 
http://windowslive.com/RediscoverHotmail?ocid=TXT_TAGLM_WL_HM_Rediscover_Storage1_042009

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Check it out.


-
Jürgen Bosch
Johns Hopkins Bloomberg School of Public Health
Biochemistry and Molecular Biology, W8708
615 North Wolfe Street
Baltimore, MD 21205
Phone: +1-410-614-4742
Fax:  +1-410-955-3655

[ccp4bb] Link two proteins into one polypeptide

[Raji Edayathumangalam]

Hi People,

Could anyone point me to successful examples for two unrelated  
proteins that have been stitched together into one single polypeptide  
chain with flexible amino acids to create a functional chimera that  
was subsequently crystallized. I've looked up a few.


I am particularly interested in understanding all the important  
considerations while designing a flexible linker even though many of  
these factors might be case dependent might be variable, obvious and  
commonsensical. Either way, I'd like to hear what folks have to say.


Thanks very much.
Raji

[ccp4bb] Postdoctoral position available

[Stewart Turley]

Postdoctoral Position available in the Laboratory of Wim Hol
Department of Biochemistry, School of Medicine,
University of Washington, Seattle, USA

Structural Biology and structure-based inhibitor design of the invasion
machinery of the malaria parasite.



JOB DESCRIPTION:

This project aims to unravel the structure of the invasion machinery of 
the
malaria parasite and to design inhibitors to interfere with the proper
functioning of the machinery. Such inhibitors might be the foundation for new
anti-malarials.

The malaria parasite invades two types of cells when in the human host:
hepatocytes and erythrocytes. This invasion is a fascinating and very complex
process, but some features are shared in the invasion of these two different
cell types. These include aldolase (yes, the glycolytic enzyme is moonlighting
here...) interacting with an actin filament, a specific myosin motor interacting
with the actin, a myosin-tail interacting protein (MTIP), and a number of
so-called GAP proteins. We have elucidated several structures of interacting
proteins in this system. There are also exciting invasion-blocking compounds
which we try to co-crystallize with their target protein.

The aims for the project in the next few years are; (i) to obtain 
additional
structural information about additional proteins and multi-protein complexes of
the machinery; and, (ii)
to assist in structure-based design of invasion inhibitors.

The project is an interdisciplinary collaboration with molecular 
modeling,
chemical synthesis, parasitolology and biochemical groups, in the Biomolecular
Structure Center of the University of Washington, Seattle, and Drexel
University, Philadelphia.

The successful candidate will have the opportunity to: (i) carry out 
protein
expression and protein chemistry studies to obtain insight into protein-protein
interactions involving the invasion machinery of the malaria parasite; (ii)
determine high resolution crystal structures of individual proteins as well as
multi-protein sub-complexes of the machinery; (iii) solve crystal structures of
invasion proteins in complex with small molecule inhibitors. There will also be
opportunities to carry out biophysical binding assays. Several expression
systems are already available for preparing soluble proteins from the invasion
machinery.

For further information regarding our studies on the invasion machinery 
see:

Bosch, J., Turley, S., Daly, T. M., Bogh, S. M., Villasimil, M. L., Roach, C.,
Zhou, N., Morrisey, J. M., Vaidya, A. B., Bergman, L. W. & Hol, W. G. J. (2006).
Structure of the Plasmodium MTIP-MyoA complex, a key component of the malaria
parasite invasion motor. Proc. Natl. Acad. Sci. USA 103, 4852-4857.

Bosch, J., Buscaglia, C. A., Krumm, B. E., Ingason, B., Lucas, R., Roach, C.,
Cardozo, T., Nussenzweig, V. & Hol, W. G. (2007). Aldolase provides an unusual
binding site for TRAP in the invasion machinery of the malaria parasite. Proc.
Natl. Acad. Sci. USA, 104, 7015-8020.
Bosch, J., Turley, S., Roach, C., Daly, T. M., Bergman, L. W. & Hol, W. G.
(2007) The Closed MTIP-MyosinA-tail Complex from the Malaria Parasite Invasion
Machinery. J Mol Biol. 372, 77-88.

And for information regarding the research in our lab see the websites:
http://www.bmsc.washington.edu/WimHol/
http://depts.washington.edu/biowww/faculty/hol.html

JOB REQUIREMENTS:  - Experience with:
- protein expression and purification methods
- characterization of purified soluble proteins
- protein crystallization
- At least three years of experience with protein structure determination
methods, including:
X-ray diffraction data collection and data processing; experimental and
molecular replacement phasing methods; density modification; crystallographic
refinement.
- Excellent interpersonal skills to function optimally in the malaria invasion
project team and to cooperate with collaborators in other groups and
institutions.


Experience in one or more of the following fields would be a plus:
- Molecular biology for protein overexpression
- Purification and characterization of multi-protein complexes
- Biophysical binding assays
- Molecular modeling for inhibitor design.


START DATE: Immediately

INSTITUTION:Department of Biochemistry
Biomolecular Structure Center
School of Medicine
Box 357742
University of Washington
Seattle, WA, 98195  USA

Please send your CV, including a description of your experience, a list of
publications, and names and email addresses of three references able to assess
your scientific experience and capabilities to: wg...@u.washington.edu

Re: [ccp4bb] Identifying an unknown ligand

[Artem Evdokimov]

Looks like oxalate with two water molecules nearby. Oxalate is a fairly
common product of PEG oxidation.

Artem

---
When the Weasel comes to give New Year's greetings to the Chickens no good
intentions are in his mind.
-Original Message-
From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of
Abhinav Kumar
Sent: Thursday, April 02, 2009 1:51 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Identifying an unknown ligand

Some more info about this structure:
Crystallization conditions: Glycerol, 0.1700M NaOAc, 25.5000% PEG-4000, 
0.1M TRIS pH 8.5

This does not sit on any crystalllographic symmetry axis.
But it sits right between two monomers and the NCS 2-fold axis.

The protein is chemotaxis protein CheX.

Environment: Some main chain amide Ns around it, as well as phobic side 
chains.

No peak in anomalous difference fourier map.


Thanks 
Abhinav 

Stanford Synchrotron Radiation Laboratory 
Joint Center for Structural Genomics 
Mail Stop 99 
Phone: (650) 926-2992 
Fax: (650) 926-3292 



Abhinav Kumar wrote:
> Hi,
>
> I am refining a structure and have a region of unmodeled density into 
> which I am trying to fit a ligand. The identity of the ligand is not 
> obvious, so I modeled a bunch of dummy atoms into the density.
> Could you please have a look at the map and pdb files and help me 
> identify this ligand?
>
> Please see http://smb.slac.stanford.edu/~abhinavk/UNL/unl.html
>
> Thanks Abhinav
> Stanford Synchrotron Radiation Laboratory Joint Center for Structural 
> Genomics Mail Stop 99 Phone: (650) 926-2992 Fax: (650) 926-3292 

Re: [ccp4bb] Eleven plausible phasing elements remain unused

[Jacob Corn]

I only deposited the high-res apo coordinates, but pdb code 2AU3 was solved
from a thulium soaked crystal. In fact, I also used dysprosium to phase
primase from a different bacterium. Go team lanthanide!

On Wed, 1 Apr 2009 15:21:16 +0100, Thomas Womack 
wrote:

>A perusal of the PDB reveals that the game of Periodic Table bingo still
>has eleven rounds to run:
>
>scandium, titanium, germanium, zirconium, niobium, neodymium,
>dysprosium, thulium, hafnium, bismuth and thorium remain absent from PDB
>entries.
...
> There
>must somewhere be a protein with a site that cries out for ThCl2(2+), an
>unexpectedly water-stable cation.

Re: [ccp4bb] friction or releasing pdb's

[Takanori MATSUURA]

Hi all,

You can find our release policy in the chapter 2 of "Annotation 
Policies" document which is available from the following URL.

http://www.wwpdb.org/docs.html

We encourage all the PDB depositors to carefully check the journal 
policy for the availability of coordinates (and experimental data)
before depositing coordinate to wwPDB or submitting papers to the 
journal.


For example, Nature says:
  the author must authorize Protein DataBank (PDB) release on the
  Wednesday of (or before) online or print publication.
From http://www.nature.com/authors/editorial_policies/availability.html
This means the PDB entry must be released BEFORE the journal 
publication.



We receive publication dates and citation information from some 
journals.  In this case, we send email that "your entry will be 
released by the journal request." to the contact person for the PDB 
entry.


For other journals, we search the publication information with using 
the deposited author names and deposited article title.  In this 
case, we send email notification to the contact person for the PDB 
entry.


In each case, depositors can request to postpone the release against 
the journal policy.  It is not the wwPDB responsibility to force to 
release data according to a journal policy.  It is the depositor's 
responsibility to follow the journal policies regarding release of 
the coordinates.  If depositors will be in any trouble with the 
journal office regarding their own decision, it's on their head.


The citation information that is sent to us from the depositors or 
community is also greatly appreciated.



Best regards,
Takanori
Annotators' leader, PDBj


From: Engin Ozkan 
Subject: Re: [ccp4bb] friction or releasing pdb's
Date: Thu, 2 Apr 2009 09:56:20 -0700


I do not mind PDB releasing only on Wednesdays at a certain time of
the day. What should happen is the authors/publishers should let PDB
know the publication date before it becomes available (supposedly 
days

or weeks before the actual publication, online or in print), and PDB
can automatically release pdb/sf the week of the publication or the
Wednesday before.

With requesting pdb releases, I take two routes: ask the journal and
ask RCSB.  Asking journals used to work, although my last attempt 
was

unsuccessful. If the journal agrees to do anything, they may let the
authors or PDB know, so after a while, the release happens. Asking
RCSB works if they can find the reference themselves, except in the
bizarre case I cited.

But still, the reader should not have to search for data supposed to
publish simultaneously with the article, right?

Engin

On 4/2/09 8:53 AM, Phil Jeffrey wrote:

PDB seem to take about a week to release coordinates that are HPUB
(hold for publication) from when we ask them to.  Sometimes they 
drop

the ball, but mostly this is what we see.  If I ask them to release
the coordinates early to get around this lag, I can largely 
guarantee

that I'll forget to update the JRNL reference.

I believe that the lead time is unnecessarily long.  Perhaps one 
day

might be more reasonable.

Phil Jeffrey

Engin Ozkan wrote:
I agree that the pdb deposition process has gotten better, but I 
still

regularly have issues with releasing of newly published
structures. There seem to be delays; just as you are reading this
brand new, interesting structure, you realize that the pdb and
structure factors are not released. Quite annoying. This is in 
large

part the fault of the authors, but maybe PDB could do better, too.



--
MATSUURA Takanori, Ph.D.
Visiting Associate Professor
Annotators' Leader, Group of Primary Annotation,
Protein Data Bank Japan (PDBj)
Research Center for Structural and Functional Proteomics,
Institute for Protein Research, Osaka University
3-2 Yamadaoka, Suita, Osaka 565-0871, JAPAN
Tel: +81-6-6879-8634
Fax: +81-6-6879-8636