Re: [ccp4bb] CCP4BB vs COVID19

[Chandra]

hi

There are developments in cyclic and constrained peptides

see for example J Am Chem Soc. 
2019 Mar 
13;141(10):4167-4181.


hope it helps


On 22/3/2020 8:01 pm, Abhishek Anan wrote:

Dear all,

Not directly related to the discussion but does anyone know of a
antiretroviral protease inhibitor that is a peptide? I am new to the
topic and read that peptides suffer from bioavailability issues. Are
there any workarounds?

best,
Abhishek


On 3/21/20, Rigden, Dan  wrote:

Hi James


5o32I is not a homolog of ORF8 - the BLAST e-value is insignificant. In
fact, rather than the EGF-like fold of 5o32I, ORF8 has an Ig-like fold
similar to ORF7 (for which there is a structure; 1xak).


https://toolkit.tuebingen.mpg.de/jobs/2717885_1


I must say I got quite excited seeing that until I noticed this pre-print
which tells the whole story very nicely, including that key position 84


https://www.biorxiv.org/content/10.1101/2020.03.04.977736v1


Best wishes

Dan


From: CCP4 bulletin board  on behalf of Patrick Shaw
Stewart 
Sent: 21 March 2020 15:41:17
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] CCP4BB vs COVID19


James, this isn't conventional structural biology, but may be of interest,
and I haven't been able get any mainstream virologists to think about it.

The protein sequences are obviously of interest, but so are the RNA
sequences at both ends of the Covid genome, which have conserved secondary
structure.  A few years ago a paper came out suggesting that wild-type
influenza has multiple "RNA thermometers", which may play an important role
in the tropism of influenza.  Similar mechanisms may exist in other
respiratory viruses, including Covid.

My take on this, and the relevant papers, are below.

Good luck to everyone and stay well,

Patrick


https://oldwivesandvirologists.blog/Covid-19-and-the-trade-off-model-of-selection/

My paper in Medical Hypotheses
http://douglas.co.uk/f_ftp1/ShawStewart_final_1-s2.pdf

Narberhaus, Franz, Torsten Waldminghaus, and Saheli Chowdhury. "RNA
thermometers." FEMS microbiology reviews 30.1 (2006): 3-16.

Chursov, Andrey, et al. "Specific temperature-induced perturbations of
secondary mRNA structures are associated with the cold-adapted
temperature-sensitive phenotype of influenza A virus." RNA biology 9.10
(2012): 1266-1274.

Yang, Dong, and Julian L. Leibowitz. "The structure and functions of
coronavirus genomic 3′ and 5′ ends." Virus research 206 (2015): 120-133.



On Fri, Mar 20, 2020 at 10:59 PM James Holton
mailto:jmhol...@lbl.gov>> wrote:
You might think that as a structural biologist you won't be able to do
much about COVID-19 anytime soon, but that is not true.  Yes, real-world
therapeutics and vaccines take time, but we have already seen just how
fast we can get started.  There are 21 PDBs already and some even have
bound ligands.  Good job Frank et al. BTW!  And my personal thanks to
all of you out there who are already hard at work on this.

I believe this forum is an ideal place to share information and ideas on
the structural biology of SARS-CoV-2 as we move forward. It's a big
virus, but there are not that many proteins in it.  If all of us
independently do the same bioinformatics and literature searches and end
up trying exactly the same thing in every lab all over the world, then
that would be more than unfortunate.  To that end, I am personally
interested on ORF8 for reasons I will go into below.  Has anyone tried
to solve it yet?  What happened?  Didn't express? Bad diffraction?
What?  Do tell.

Some of us, as you may have heard, are stuck at home, our beamlines and
labs dark while we shelter-in-place.  That doesn't mean our hands are
tied.  We are still allowed to think. The fraction of the human race
that has a snowball's chance in Hades of figuring out this bug is very
very small.  Structure may be your main skill set, but you are still a
biologist.  Do you know how to run a PCR machine?  Do you know how to
pipette?  You might think that anybody can do it, but that is really not
the case. Ever trained a new student on sterile technique?  How many
days did that take?  Now remember that your student was no dummy and
already studying biology.  Everyone reading this will make an excellent
volenteer at the very least.  I'm not saying this to belittle the
average human, only to say that we scientists, moving in the circles we
do, often forget that we have uncommon capabilities.

For example, I also believe we can be useful in assay development. The
void left by the dearth and delay of test results has been filled with
fear, and that is a big problem.  The tests, as defined, are
straightforward, but also extremely regimented like any good laboratory
protocol should be.  The US CDC's instructions for academic labs are here:
https://www.cdc.gov/coronavirus/2019-nCoV/lab/index.html
My question is: how can this test be made faster, using more commonplace
supplies, in 

[ccp4bb] Bidentate GLU vs two monodentate GLU in metalloproteins

[Chandra Prakash Tiwari]

Dear all,
What is more preferable or conserved by evolution of lifeform point of view
between glutamate acting as bidentate ligand by its carboxylate group or
two monodentate GLU at metal binding site in natural metalloenzymes.
I was thinking 2 GLU are better than one bidentate GLU because if one GLU
gets mutated other GLU will still be able to form a coordination bond to a
metal.
Please comment and help if i am right or wrong. metal is Fe2+. In short
which will evolve with time a bidentate GLU or 2 monodentate GLU.



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

[ccp4bb] Bidentate GLU vs two monodentate GLU in metalloproteins

[Chandra Prakash Tiwari]

Dear all,
What is more preferable or conserved by evolution point of view between
glutamate acting as bidentate ligand by its carboxylate group or two
monodentate GLU at metal binding site in natural metalloprotein.
I was thinking 2 GLU are better than one bidentate GLU because if one GLU
gets mutated other GLU will still be able to form a coordination bond to a
metal.
Please comment and help if i am right or wrong.



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

[ccp4bb] help

[Chandra]

Hello all

Does anyone knows of a review that highlights the first examples of the 
use of NMR combined with X-ray crystallography to solve the structures 
of proteins


thanks

chandra

Re: [ccp4bb] Modelling protein/protein interfaces

[Chandra]

haddock may be an appropriate tool

https://haddock.science.uu.nl/



On 26/1/2018 8:34 PM, Thomas Krey wrote:


Dear colleagues,

Is there any appropriate tool for docking an interacting protein to a 
relatively large protein-protein interface (>2500 A2). We are facing a 
hetero-oligomer for which we approximately know the interfaces as well 
as the structures of the individual protomers and would like to model 
the hetero-oligomer. I have done some small molecule docking before, 
but the size of the interface seem to be prohibitive for using the 
same tools in the case of such a protein-protein interaction.


Any help or suggestion would be highly appreciated.

Best wishes

Thomas

Re: [ccp4bb] Predict effects of mutations on protein stability and protein-protein interaction

[Chandra]

duet is quite good too

http://biosig.unimelb.edu.au/duet/


On 25/11/2017 1:20 AM, Vivoli, Mirella wrote:

Dear Wenhe,
I have used iMutant 2.0.
I hope it helps.
Below the link of a paper related to the prediction server:
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1160136/

Best,

Mirella

Get Outlook for Android 


*From:* CCP4 bulletin board  on behalf of WENHE 
ZHONG 

*Sent:* Friday, November 24, 2017 6:08:28 PM
*To:* CCP4BB@JISCMAIL.AC.UK
*Subject:* [ccp4bb] Predict effects of mutations on protein stability 
and protein-protein interaction

Dear Community,

A little bit out-of-topic here. We have a few interesting sites would 
like to mutate on proteins to test the protein stability and improve 
the protein-protein interactions. Before moving forward to the 
site mutagenesis experiment, we like to do some prediction first to 
narrow down the candidates. We only know that SDM is good server to 
predict the protein stability. Do you have other servers or softwares 
can recommend? We can do a cross comparison.


Thanks.

Kind regards,
Wenhe

[ccp4bb] AutoSol invalid MTZ column_types error

[Mintu Chandra]

Dear all,


I am getting an error message "Invalid MTZ column_types for the given
miller_array." while I try to run AutoSol. The last line in the log is
"Adding array ['N(+)', 'N(-)'] to output file with type II".
Do you have any suggestion how to fix this error?

Thanks a lot in advance. Best,

Mintu

[ccp4bb] relative domain motion calculation...

[Mintu Chandra]

Dear All,
I am working with a protein containing tandem domains, connected with
3-4 residue linker. I want to calculate the relative
motion/orientation of one domain with respect to the hinge region as
well as with respect to the other domain. Both the domains have very
less seq. identity (20%). Please suggest some web server to calculate
the same.

Thanks,

Mintu

[ccp4bb] protocol for seleno-methionine incorporation...

[Mintu Chandra]

Dear all,
Is there any standard protocol to know the number of incorporated
seleno-methionine in a protein ??? I am growing one of the protein in
minimal media for phase determination.
I am using mass spectrometry facility to know the incorporated
seleno-methionine but I wanted to know whether I should use the purified
protein  or  digested protein for mass spectrometry.

Thanks,

Mintu Chandra.

-- 
Mintu Chandra
Senior Research Fellow
IISER BHOPAL
mi...@iiserb.ac.in
+918085288853

[ccp4bb] Align linux version

[Mintu Chandra]

Dear CCP4 users,

We are trying to align two different structures. It would be helpful
for us if anybody has linux compiled align version?

Regards,

Mintu Chandra

Re: [ccp4bb] off topic: rmsf in simulation

[Chandra Verma]

to complement the very nice description by jeremy, you may wish to try  
and decompose the vibrational modes to get this sense by focussing on  
the origins of  the red shift in the vibrational spectrum and this  
accounts largely for the increased vibrational entropies upon  
complexation. This paper may be used as a guide
(Dissecting the vibrational entropy change on protein/ligand binding:  
burial of a water molecule in bovine pancreatic trypsin inhibitor J  
Phys Chem B  2001 105 8050-8055)



There is some very nice work by Olano  Rick in

JACS 2004 126:7991 on Hydration free energies and entropies for water  
in protein interiors.


and by carol post on how the increased entropies upon complexation are  
the origin of the mechanism of some drugs. (for example Influence of  
an Antiviral Compound on the Temperature Dependence of Viral Protein  
Flexibility and Packing: a Molecular Dynamics Study J. Mol. Biol.  
(1998) 276: 331-337)




Quoting Jeremy Tame jt...@tsurumi.yokohama-cu.ac.jp:

Different proteins do different things. Some adopt fewer  
conformations and a more rigid structure after binding
a ligand, and others do the opposite. Haemoglobin is a nice example  
of a protein that becomes a lot more flexible
after picking up ligands. For any reaction of the kind P + L - PL  
there is an entropy cost of making one molecule
from two. For the protein to activate low frequency modes in the  
complex is one way to compensate for this by
increasing the entropy of the bound form. The paper by Sturtevant  
(PNAS 74, 2236, 1977) is worth a read, as is
Cooper and Dryden (Eur Biophys J, 11, 103, 1984), if you are  
interested in relating fluctuations to thermodynamics.
All too often people attempt direct comparisons of structural models  
and affinities without realising that the so-called
angstroms to calories problem often frames the question in a form  
that cannot be answered sensibly. For
example, imagine a protease which is produced as a zymogen. Both  
forms may have essentially identical crystal
structures even though the zymogen is more flexible. The protease  
can be activated by loss of vibrational modes
in the unbound state which are re-awakened in the complex with  
substrate; hence the zymogen will have lower
substrate binding and activity. You might be interested in a review  
by Homans (ChemBioChem 6, 1585, 2005) which
discusses the use of NMR to look at entropy changes in  
protein-ligand binding reactions. It is by no means unusual
for a residue's entropy to increase in the bound state, although in  
your case it seems to be the whole protein!


On Dec 9, 2012, at 1:05 PM, anita p wrote:


Hi All,
I am trying to understand the mechanism of protein-peptide  
interaction in two complexes (protein-pepA and protein-pepB).
While trying to perform some simulation experiments, I find that  
the root mean square fluctuation (RMSF) by residues of protein in  
the complex is higher than that of the protein alone.
Please refer the figure attached to this email. pepA binds with  
higher affinity (in uM-range) than pepB according to invitro studies.


Does this happen normally?? Please advice.
Thanks in advance
Anita
RMSF.png

Re: [ccp4bb] Protein-Protein Interactions

[Manish Chandra Pathak]

In addition to what Francisco suggested, looking at the sequences with 
evolutionary highlited residues will provide additional info.

If modeled structures  are available , which is not the case with this query, 
investigating the corresponding electrostat potential using APBS  might give 
evidence to cross check results obtained from other method. 

Hope it helps
Manish


Manish Chandra Pathak, Ph.D. 
Indian Institute of Science Education and Research 
ITI (Gas Rahat) Building Govindpura, 
Bhopal 462023 India 
tel: +91-750-4092340


--
On Thu, Aug 2, 2012 1:28 PM EDT Francisco Hernandez-Guzman wrote:

Hi Lorenzo,

I forgot to add that any experimental data that you can provide to guide the 
modeling is highly recommended and often necessary to validate your 
predictions. Modeling can be quite useful but you should be aware of its 
strengths and weaknesses.

Cheers,

Francisco

From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of 
Francisco Hernandez-Guzman
Sent: Thursday, August 02, 2012 10:21 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Protein-Protein Interactions

Hi Lorenzo,

If the structure for your receptor is unknown, then you can use Homology 
Modeling methods to get a rough idea of the structure, MODELLER is a well know 
tool for this (http://salilab.org/modeller/). Of course depending on your % 
similarity to the template, the higher the % similarity, the more reliable 
your structure may be (of course assuming there are no major conformational 
changes, etc.)

Now, to figure out the sites of interaction, you could use a shape based 
complementarity approach like the one used in the ZDOCK algorithm 
(http://zdock.umassmed.edu/software/). This gets to be a little bit trickier 
if your % similarity to your template is low, because the dissimilarity is 
often due to surface residue differences, which are obviously the ones you're 
interested on. On the other hand, if the source of interaction is driven 
mainly by hydrophobic forces, then an analysis using the spatial aggregation 
propensity method 
(http://pubs.acs.org/doi/abs/10.1021/jp911706q?journalCode=jpcbfk) may reveal 
interesting sites of aggregation. This method is a little bit more forgiving 
that the shape complementarity one because of the intrinsic averaging that 
goes on to determine the site of aggregation.

All of these methods and other simulations tools are available in the 
Discovery Studio suite from Accelrys.

Disclaimer: I work for Accelrys as their Product Manager for the Life Science 
Modeling and Simulations suite of products. So, if you're interested in 
evaluating and gain access to these tools please contact me directly.

Kind regards,

Francisco
Sr. Product Manager
http://accelrys.com

From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Dr. 
Lorenzo Finci
Sent: Thursday, August 02, 2012 6:07 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] Protein-Protein Interactions

Dear Colleagues,

I have a question for all of you bioinformatics oriented structural 
biologists: How do I predict the sites of protein-protein interactions between 
two receptors that have been proven to interact biochemically but lack 
specific details regarding proximity. This is not a straightforward question 
for me, and I believe it is somewhat complicated. The complicated scenario 
involves a multitude of different subunits and isoforms. Also, there is not 
structural data to support all components involved, and thus I presume I 
should use the sequence based software. I am aware that there are different 
types of prediction software, either sequence or structure based predictions 
using different algorithms:
http://rosettadesigngroup.com/blog/58/10-protein-protein-interface-prediction-servers/

Receptor 1:
-Has 5 predicted subunits (Alpha)2-(Beta)2-(Gamma)1
1. Alpha (6 isoforms)
2. Beta (3 Isoforms)
3. Gamma (3 Isoforms)

Receptor 2:
-Is believed to be composed of (Alpha)3-(Beta)2
1. Alpha (4 isoforms)
2. Beta(1 isoform)

Any advice or recommendation will be well appreciated!

Sincerely,
lorenzo
Lorenzo Ihsan FInci, Ph.D.
Postdoctoral Scientist, Wang Laboratory
Harvard Medical School
Dana-Farber Cancer Institute
Boston, MA
Peking University
The College of Life Sciences
Beijing, China

Re: [ccp4bb] modeling thioester bond

[Manish Chandra Pathak]

Hi Sebastiano, 


In this case, having your own dictionary for both residues together will be 
useful. Same dictionary will be used in refmac refinement too, if you need 
precise refined positions. 


It will be easier, if you do the editing in text file. Get the CIF files for 
Cys-X together, rename residues and satisfy the valency - bond length etc. Let 
me know if you need help.

all the best
Manish






 From: Sebastiano Pasqualato sebastiano.pasqual...@gmail.com
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Thursday, July 26, 2012 6:06 PM
Subject: Re: [ccp4bb] modeling thioester bond
 



Thanks Manish.
With the help of Paul and using JLigand, which is nicely connected to coot in 
version 0.7-pre1, I managed to create the bond and have the correct library 
CIF file.


Still, I can't manage to regularize the bond and residues close to it. It 
looks like I have to go through refcmac via structure idealisation.


Unlike in the case of sphere refinement, when you have a density map, in my 
case I don't know how to regularize a zone which is comprised between two 
different chains.


Thanks for the tip,
ciao,
s



On Jul 26, 2012, at 2:19 PM, Manish Chandra Pathak wrote:



Just defining LINK in your pdb file should work. Try this format.
LINK NZ  LYS A 233 C4A PLP A 400 


OR

You need a dictionary file to tell the COOT that the two residues are 

connected. If an instance already exist in PDB, then you can download 

the refmac dictionary from  http://xray.bmc.uu.se/hicup/
Otherwise, use PRODRG Dundee server with coordinates of both 

residues (Cys-X) to create a new residue dictionary. 

all the best
Manish



Manish Chandra Pathak, Ph.D.
Indian Institute of Science Education and Research
ITI (Gas Rahat) Building
Govindpura, Bhopal 462023  India
tel: +91-750-4092340







From: Sebastiano Pasqualato sebastiano.pasqual...@gmail.com

To: CCP4BB@JISCMAIL.AC.UK 

Sent: Wednesday, July 25, 2012 9:48 PM

Subject: [ccp4bb] modeling thioester bond









No answers yet from the COOTbb (yet), so I'm cross posting this to the ccp4bb.

Sorry for the double post.













Hi there,

I'd like to model a thioester bond, starting from protein A with an exposed 
Cys, and having juxtaposed the carboxyl group of a C-terminal residue of 
protein B.





The way I would like to proceed is by:





1) telling coot that there's a bond between the S atom and the C atom of the 
two groups;

2) regularize that zone in order to have something that makes chemical sense.





Could you advice me on how to proceed for the two steps?

That is, how do I tell coot that the bond exists and it has to be taken into 
account?





Thanks in advance,

ciao,

s











-- 

Sebastiano Pasqualato, PhD



Crystallography Unit

Department of Experimental Oncology

European Institute of OncologyIFOM-IEO Campusvia Adamello, 1620139 - Milano

Italy



tel +39 02 9437 5167

fax +39 02 9437 5990



please note the change in email address!sebastiano.pasqual...@ieo.eu




















-- 
Sebastiano Pasqualato, PhD

Crystallography Unit
Department of Experimental Oncology
European Institute of OncologyIFOM-IEO Campusvia Adamello, 1620139 - Milano
Italy

tel +39 02 9437 5167
fax +39 02 9437 5990

please note the change in email address!sebastiano.pasqual...@ieo.eu

Re: [ccp4bb] Self-rotation Patterson maps

[Manish Chandra Pathak]

Hello John, 

In my struggle with self-rotation function, I found this extremely useful to 
start with. Specially the fig 4 on page 107. 


Characterizing a Crystal From an Initial Native Dataset by Michael R. Sawaya
http://www.springerlink.com/content/k024q83484477j42/

The peaks in the log files are more helpful than the peaks in Patterson map 
itself.


all the best
Manish


From: Ian Tickle ianj...@gmail.com
To: CCP4BB@JISCMAIL.AC.UK
Sent: Monday, August 29, 2011 5:25 PM
Subject: Re: [ccp4bb] Self-rotation Patterson maps


Hi John,

There's some by now fairly ancient tutorial material on the SRF (though of 
course the theory hasn't changed) here:

http://www.ccp4.ac.uk/dist/ccp4i/help/modules/appendices/mr-bathtutorial/mrbath2.html#section2.8

(and the following section 2.9) and here:

http://www.ccp4.ac.uk/dist/ccp4i/help/modules/appendices/mr-bathtutorial/mrbath5.html

(see section 5.2).

Cheers

-- Ian


On Mon, Aug 29, 2011 at 7:12 AM, john peter jpeter1...@gmail.com wrote:

Hello,

 I wish to learn about self-rotation Patterson maps such as how to
interpret a map and find out non-crystallographic symmetry. Could you
suggest some good papers (for dummies), web-sites etc.

Thanks a lot for your help.

John

[ccp4bb] Post Doc Opportunity to post, please

[Chandra Farnsworth]

I would like to have the following Post Doc Research Fellow opportunity
posted please.
Thanks very much,
Chandra
*Chandra Farnsworth*
*Staffing Consultant*

Genentech, Inc.
Research and Early Development
650-225-7110
---
This position is in the Daniel Kirchhofer Lab
http://www.gene.com/gene/research/postdoctoral/mentors/strchembio/kirchhofer/




Description:

We are looking for a Postdoctoral Fellow to study the structural and
functional basis of how antibodies regulate catalysis of proteases. Another
aspect of the project is the biochemical and biological characterization of
a novel trypsin-like serine protease involved in inflammatory disease.



Requirements:

The candidate should have a Ph.D. in biochemistry or biophysics or related
field with a strong background in protein chemistry. Candidates with
expertise in protein purification/characterization and

 knowledge of protein structures (incl. Pymol) are preferred. The individual
should be highly motivated, demonstrate creative thinking, able to work
independently and interact well with other members of the lab.



http://www.gene.com/gene/careers/

Re: [ccp4bb] Solvent accessible regions of the channel (Pore)

[Manish Chandra Pathak]

Hi Bashir, 

In Pymol:
1. Show---Surface (will generate surface around the protein)
2. Setting-Surface---Cavities  Pockets Only. (will show only Cavities 
inside)
3.  Use cavity_cull command to set the filter for cavities. 
(eg. set cavity_cull, 40)

Manish


--
Manish Chandra Pathak, Ph.D.
Department of Biochemistry
Emory University School of Medicine
1510 Clifton Road, NE, Room G235
Atlanta, GA  30322  USA
Tel: +1-404-727-2563 Fax: +1-404-727-2738




- Original Message 
 From: Muhammed bashir Khan muhammad.bashir.k...@univie.ac.at
 To: CCP4BB@JISCMAIL.AC.UK
 Sent: Wed, April 7, 2010 9:58:20 AM
 Subject: [ccp4bb] Solvent accessible regions of the channel (Pore)
 
 Dear all;

Can anybody help me how I can make the solvent accessible 
 surface(inside)
of the channel(pore)by pymol, or by any other 
 programme.

Thanks in Advance

-- 
Muhammad Bashir 
 Khan
Department for Biomolecular Structural Chemistry
Max F. Perutz 
 Laboratories
University of Vienna
Campus Vienna Biocenter 5
A-1030 
 Vienna
Austria

Phone: +43(1)427752224
Fax: +43(1)42779522


  

Re: [ccp4bb] How to compare the binding affinity between two domains structurally?

[Chandra Verma]

if the link between the domains is not part of the interface then perhaps
a good first approximation may be to cut the linker and then use a
program such as APBS to compute the binding energy..


 Hi,

 Recently we determined two structures of the same protein in complex with
 different molecules.  The protein contains two domains (called domain A
 and
 B here).  In the two structures, domain A and B have different
 arrangements
 relative to each other, resulting different interaction interface.  I want
 to know which inter-domain interaction is stronger.   Is there a way to
 quantatively compare the interaction energy or intensity between the two
 domains?  I have calculated the buried surface area.  However, just
 comparing the buried surface does not provide definitive answer, given
 that
 the interacting residues on the interface are also different.

 BTW, we were not able to purify individual domains, so we cannot measure
 the
 binding affinity by wet lab approaches (so far).

 Thank you in advance for your inputs.

 --
 Best regards,

 Joe

Re: [ccp4bb] How to fix sidechain rotamers for Refmac?

[Manish Chandra Pathak]

Hi Peter, 

I think, instead of tightening the weighing term, giving it slight freedom 
would have helped.
If data quality is good at 1.8 ang, then it certainly deserves more freedom. 
What do you think. 

best wishes
Manish



Manish Chandra Pathak, Ph.D.
Department of Biochemistry
Emory University School of Medicine
1510 Clifton Road, NE, Room G235
Atlanta, GA  30322  USA
Tel: +1-404-727-2563 Fax: +1-404-727-2738
email: mcpa...@emory.edu








From: Peter J Stogios p.stog...@utoronto.ca
To: CCP4BB@JISCMAIL.AC.UK
Sent: Wednesday, September 30, 2009 12:22:54 PM
Subject: [ccp4bb] How to fix sidechain rotamers for Refmac?

Hi,

I'm working with a 1.8 A structure with Coot and Refmac, and there are many 
sidechain rotamers that show very clear difference density peaks for setting 
their correct positions.  However, Refmac continuously moves the rotamers back 
into negative density peaks.  It's really quite silly because often there is an 
obvious positive density peak near to a negative density peak.

I have tried using automatic geometry weighting and manually setting the 
weighting term to a very tight 0.025, but each has no effect.  I have also 
tried increasing the torsion angle restraint term to 2.0 but this also has no 
effect.

Does anyone have any suggestions?  Is there any way to fix atom positions for 
Refmac?

Thanks in advance,

~
Peter J Stogios, Ph.D.
Postdoctoral Research Fellow
e: p.stog...@utoronto.ca
p: 416-978-4033
w: http://www.uhnres.utoronto.ca/centres/proteomics/

Structural Proteomics in Toronto Research Group, University Health Network
C.H. Best Institute
112 College Street, Room 70
Toronto, Ontario, Canada M5G 1L6



  

Re: [ccp4bb] Ligand binding in multiple conformation

[Chandra Verma]

An extension of this topic: does anyone know of examples where for 
complexes between the two proteins A and B, protein A has two different 
conformations at its interface?


thanks

chandra




-Original Message-
From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of
Aaron Oakley
Sent: Tuesday, January 13, 2009 1:19 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Ligand binding in multiple conformation

 

  

I had a question about flexibility in ligand binding in an enzyme


active site.
  

Is it possible for a substrate/product analogue to bind in more than


one conformation in the active site.

Yes. It is even possible for portions of a ligand to be disordered and
not discernable in electron density.

  

Since the ligand/enzyme interactions are very specific I am a little


confused about this.
  

Also which program would you use if you have to refine with alternate


ligand conformation.

REFMAC will do this just fine. Just make sure you have the A, B
conformers set in the PDB file.

EG:

ATOM 72  CA APRO A   7  -2.619  28.983  -0.796  0.62  6.48
C  
ATOM 73  CA BPRO A   7  -2.226  29.044  -0.847  0.38  5.76
C  

  

Re: [ccp4bb] Ligand binding in multiple conformation

[Chandra Verma]

hi

yes this may happen in some circumstanceshave a look at

J Am Chem Soc. 2008 Oct 15;130(41):13514-5.


 Hi,

 I had a question about flexibility in ligand binding in an enzyme active
 site.  Is it possible for a substrate/product analogue to bind in more
 than
 one conformation in the active site.  Since the ligand/enzyme interactions
 are very specific I am a little confused about this.
 Also which program would you use if you have to refine with alternate
 ligand
 conformation.
 Please mention if you have ever come across any paper that explains such a
 phenomena.
 Thanks a lot.

 Mariah
 --
 Mariah Jones
 Department of Biochemistry
 University of Florida

Re: [ccp4bb] Setting drops with proteins that are hard to concentrate

[Manish Chandra Pathak]

Dear Matt, 

In one of our project, the protein of interest used to elute from Ni-column 
with 
~20 other proteins. I also guessed it due to surface Cysteines. 

Addition of 2mM free Cys in the protein sample helped, which competed with 
free Cys on the protein surface and, ultimately, gave happy single band on 
sds-page.

All the best
Manish



--
Center for Advanced Research in Biotechnology 
University of Maryland Biotechnology Institute 
9600 Gudelsky Drive, Rockville 
MD 20850  USA 
Tel: +1-240-314-6130;  fax: +1-240-314-6255


- Original Message 
From: Bottomley, Matthew [EMAIL PROTECTED]
To: CCP4BB@JISCMAIL.AC.UK
Sent: Thursday, April 24, 2008 7:35:46 AM
Subject: [ccp4bb] Setting drops with proteins that are hard to concentrate

Setting drops with proteins that are hard to concentrate  Dear All,
I have a 50kDa protein that is soluble and monodisperse at up to approx 1mg/ml 
(after Ni-affinity and size-exclusion chromatography).
However, it aggregates (probably both via disulphides and via 
'sticky/hydrophobic patches') when I concentrate it towards 2-3mg/ml, even in 
the presence of several detergents. I don't want to add DTT since my protein 
should have several intramolecular disulphidesalthough I do have 2 free 
Cysteines, partially exposed. I have already tried mutating the Cysteines, with 
little improvement.
Any suggestions for obtaining 5-10mg/ml? 
Does anybody have good experiences with usin L-Arg and L-Glu (e.g. At 50mM)  to 
aid concentrating (as in the Golovanov AP paper, JACS, 2004, pages 8933...)
Thanks for any input!
Yours,
Matt

Matthew J. Bottomley, Ph.D.
Senior Research Biochemist
IRBM / MRL-Rome








  

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Re: [ccp4bb] refinement problem of labelled RNA complex

[Manish Chandra Pathak]

Hi, 

It's much easier to begin with the cif file created by Refmac5 
itself, but this cif file needs to be checked and corrected 
manually for the restrains. Certainly, Iodine is not happy 
with the current restrains. 

Most of the time, i have found problems beginning with the 
protonation and the bond order and then extending to the 
geometrical freedom of the molecule.  

You may want to try Monomer sketcher, where you can manipulate 
the monomers visually.

all the best
Manish



-
Center for Advanced Research in Biotechnology
University of Maryland Biotechnology Institute
9600 Gudelsky Drive, Rockville
MD 20850 USA
Tel: +1-240-314-6130; fax: +1-240-314-6255





--- ºîº£·å [EMAIL PROTECTED] wrote:

 Dear all;
 I am a fresher of Refmac5. And now I'm trying to refine a protein and 
 labelled RNA complex using
 Refmac. It's a Iodo-U labelled RNA.  Unfortunately, these is no appropriate 
 lib file in the
 current refmac dictionaries. So I use cif file which created by Refmc itself, 
 surprisingly, the
 base of uridine distorted greatly. Could someone like to tell me how to make 
 the appropriate cif
 file to solve this problem. Another question is that the RNA has a phosphate 
 group in 5',  How
 to define this in the cif file?
 
 Could someone  help me? Thanks a lot!
Best wishes,

Hai-Feng Hou

 Biological macromolecule group
 Beijing Synchrotron Radiation Facility
 Institute of High Energy Physics
 Chinese Academy of Sciences
 Beijing, China
 Tel: +86-10-88234208
 E-mail: [EMAIL PROTECTED]
 
2008-02-18
 



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Re: [ccp4bb] .cif file

[Manish Chandra Pathak]

Hello Vineet,

This seems old one and doesn't have all restraints. 

you may use refmac dictionary for spermine. I am sure this 
will work in coot also. you may find it at

/ccp4-6.0.2/lib/data/monomers/s/SPM.cif 

thanks
Manish



Vineet Gaur [EMAIL PROTECTED] wrote: Hi all
 I am using COOT for model building and refinement.
 i have to introducea ligand in my model
 i have downloaded .cif file from RCSB.
 However while importing the cif file i m getting the warning message of having 
No restraints in the CIF file
 
 is there any problem in the format of the following cif file
 
 
data_SPM
# 
_chem_comp.idSPM 
_chem_comp.name  SPERMINE 
_chem_comp.type  NON-POLYMER 
_chem_comp.pdbx_type HETAI 

_chem_comp.formula   C10 H26 N4 
_chem_comp.mon_nstd_parent_comp_id   ? 
_chem_comp.pdbx_synonyms ? 
_chem_comp.pdbx_formal_charge0 

_chem_comp.pdbx_initial_date 1999-07-08 
_chem_comp.pdbx_modified_date2007-08-16 
_chem_comp.pdbx_ambiguous_flag   N 
_chem_comp.pdbx_release_status   REL 

_chem_comp.pdbx_replaced_by  ? 
_chem_comp.pdbx_replaces ? 
_chem_comp.formula_weight202.340 
_chem_comp.one_letter_code   ? 

_chem_comp.three_letter_code SPM 
_chem_comp.pdbx_model_coordinates_details? 
_chem_comp.pdbx_model_coordinates_missing_flag   N 
_chem_comp.pdbx_ideal_coordinates_details? 

_chem_comp.pdbx_ideal_coordinates_missing_flag   N 
_chem_comp.pdbx_model_coordinates_db_code1DPL 
_chem_comp.pdbx_processing_site  ? 
# 
loop_
_chem_comp_atom.comp_id 
_chem_comp_atom.atom_id 

_chem_comp_atom.alt_atom_id 
_chem_comp_atom.type_symbol 
_chem_comp_atom.charge 
_chem_comp_atom.pdbx_align 
_chem_comp_atom.pdbx_aromatic_flag 
_chem_comp_atom.pdbx_leaving_atom_flag 
_chem_comp_atom.pdbx_stereo_config 

_chem_comp_atom.model_Cartn_x 
_chem_comp_atom.model_Cartn_y 
_chem_comp_atom.model_Cartn_z 
_chem_comp_atom.pdbx_model_Cartn_x_ideal 
_chem_comp_atom.pdbx_model_Cartn_y_ideal 
_chem_comp_atom.pdbx_model_Cartn_z_ideal 

_chem_comp_atom.pdbx_ordinal 
SPM N1   N1   N 0 1 N N N -1.386 5.560  25.140 -0.292 0.012  7.961  1  
SPM C2   C2   C 0 1 N N N -2.248 4.344  25.282 0.514  -0.023 6.734  2  
SPM C3   C3   C 0 1 N N N -1.844 3.214
  24.376 -0.408 0.014  5.515  3  
SPM C4   C4   C 0 1 N N N -2.834 2.063  24.495 0.432  -0.022 4.237  4  
SPM N5   N5   N 0 1 N N N -2.313 0.871  23.720 -0.453 0.013  3.066  5  
SPM C6   C6   C 0 1 N N N -3.235 -
0.310 23.760 0.411  -0.024 1.880  6  
SPM C7   C7   C 0 1 N N N -2.669 -1.391 22.871 -0.451 0.011  0.617  7  
SPM C8   C8   C 0 1 N N N -3.306 -2.753 23.219 0.450  -0.028 -0.617 8  
SPM C9   C9   C 0 1 N N N -2.886
 -3.777 22.188 -0.412 0.006  -1.880 9  
SPM N10  N10  N 0 1 N N N -3.212 -5.172 22.585 0.453  -0.031 -3.066 10 
SPM C11  C11  C 0 1 N N N -2.084 -5.778 23.391 -0.433 0.005  -4.237 11 
SPM C12  C12  C 0 1 N N N -2.443
 -7.162 23.912 0.407  -0.032 -5.515 12 
SPM C13  C13  C 0 1 N N N -1.448 -8.158 23.367 -0.515 0.005  -6.734 13 
SPM N14  N14  N 0 1 N N N -0.553 -8.726 24.427 0.292  -0.029 -7.961 14 
SPM HN11 1HN1 H 0 0 N N N -1.660
 6.326  25.754 -0.739 0.916  7.988  15 
SPM HN12 2HN1 H 0 0 N N N -1.355 5.869  24.168 0.354  -0.014 8.735  16 
SPM H21  1H2  H 0 1 N N N -2.281 4.005  26.343 1.181  0.838  6.713  17 
SPM H22  2H2  H 0 1 N N N -3.322
 4.602  25.135 1.105  -0.939 6.715  18 
SPM H31  1H3  H 0 1 N N N -1.722 3.550  23.320 -1.074 -0.847 5.536  19 
SPM H32  2H3  H 0 1 N N N -0.796 2.883  24.565 -0.999 0.930  5.534  20 
SPM H41  1H4  H 0 1 N N N -3.061
 1.809  25.556 1.098  0.839  4.215  21 
SPM H42  2H4  H 0 1 N N N -3.862 2.355  24.178 1.023  -0.938 4.217  22 
SPM HN5  HN5  H 0 1 N N N -1.378 0.612  24.037 -0.976 -0.849 3.071  23 
SPM H61  1H6  H 0 1 N N N -3.427
 -0.667 24.798 1.078  0.838  1.889  24 
SPM H62  2H6  H 0 1 N N N -4.284 -0.042 23.493 1.002  -0.940 1.891  25 
SPM H71  1H7  H 0 1 N N N -2.780 -1.142 21.789 -1.117 -0.851 0.608  26 
SPM H72  2H7  H 0 1 N N N -1.555
 -1.427 22.916 -1.042 0.927  0.607  27 
SPM H81  1H8  H 0 1 N N N -3.068 -3.077 24.258 1.117  0.833  -0.608 28 
SPM H82  2H8  H 0 1 N N N -4.414 -2.687 23.323 1.041  -0.944 -0.607 29 
SPM H91  1H9  H 0 1 N N N -3.319
 -3.536 21.189 -1.079 -0.855 -1.889 30 
SPM H92  2H9  H 0 1 N N N -1.802 -3.675 21.945 -1.003 0.922  -1.891 31 
SPM HN0  HN0  H 0 1 N N N -3.451 -5.748 21.778 0.976  0.831  -3.071 32 
SPM H111 1H11 H 0 0 N N N -1.133
 -5.797 22.808 -1.099 -0.857 -4.215 33 
SPM H112 2H11 H 0 0 N N N -1.767 -5.101 24.219 -1.023 0.921  -4.217 34 
SPM H121 1H12 H 0 0 N N N -2.512 -7.194 25.024 1.074  0.830  -5.536 35 
SPM H122 2H12 H 0 0 N N N -3.497
 -7.443 23.683 0.998  -0.948 -5.534