[ccp4bb] off topic: Add ligands/substrate directly into expression medium?

[Liuqing Chen]

Hello everyone!
  In which condition can we add ligand/substrate directly to the expression 
medium for overexpression protein?  One protein of i crystallized is a enzyme 
that catalyze sterols substrate, as we know the solubility of sterols is very 
poor, and the Km of the enzyme towards sterols around 0.5-1mM.
Can i add sterols directly to the medium , when i expression my protein ?
If not , then in which condition can we do like this,  for inhibitor ?  i 
remeber i read one paper did like this. 

Sincerely!
Liuqing chen

Re: [ccp4bb] Non crystallography question

[amin sagar]

Seems like you are looking for something like DARA.

https://dara.embl-hamburg.de/

-Amin.
On Sat, 9 Dec 2017 at 3:21 AM, Vands  wrote:

> Hi All,
>Is there any Web server available where I can input my SAXS
> profile and it will give all closely matched PDB structures from the all
> available PDB.
>
>
> --
> Vandna Kukshal
> Postdoctral Research Associate
> Dept. Biochemistry and Molecular Biophysics
> Washington University School of Medicine
> 660 S. Euclid
> , Campus
> Box 8231
> St. Louis, MO 63110
>

Re: [ccp4bb] Non crystallography question

[Parthasarathy Sampathkumar]

Hi Vandna,

A typical approach would be to generate different models (experimental or
otherwise, if X-ray structure it could be "completed" further by modeling
missing loops / side-chains) of your sequence to calculate a SAXS profile,
and then compare / fit those to experimental SAXS profile. Some tools are
available at salilab.org

Hope this helps,
Best Wishes
Partha


On Fri, Dec 8, 2017 at 4:50 PM, Vands  wrote:

> Hi All,
>Is there any Web server available where I can input my SAXS
> profile and it will give all closely matched PDB structures from the all
> available PDB.
>
> --
> Vandna Kukshal
> Postdoctral Research Associate
> Dept. Biochemistry and Molecular Biophysics
> Washington University School of Medicine
> 660 S. Euclid
> , Campus
> Box 8231
> St. Louis, MO 63110
>

Re: [ccp4bb] Non crystallography question

[Vands]

Yes thank you very much .. I used it two years back and just blanked out.

:)
-Vandna

On Fri, Dec 8, 2017 at 3:55 PM, amin sagar  wrote:

> Seems like you are looking for something like DARA.
>
> https://dara.embl-hamburg.de/
>
> -Amin.
>
> On Sat, 9 Dec 2017 at 3:21 AM, Vands  wrote:
>
>> Hi All,
>>Is there any Web server available where I can input my SAXS
>> profile and it will give all closely matched PDB structures from the all
>> available PDB.
>>
>>
>> --
>> Vandna Kukshal
>> Postdoctral Research Associate
>> Dept. Biochemistry and Molecular Biophysics
>> Washington University School of Medicine
>> 660 S. Euclid
>> , Campus
>> Box 8231
>> St. Louis, MO 63110
>>
>


-- 
Vandna Kukshal
Postdoctral Research Associate
Dept. Biochemistry and Molecular Biophysics
Washington University School of Medicine
660 S. Euclid, Campus Box 8231
St. Louis, MO 63110

[ccp4bb] Non crystallography question

[Vands]

Hi All,
   Is there any Web server available where I can input my SAXS
profile and it will give all closely matched PDB structures from the all
available PDB.

-- 
Vandna Kukshal
Postdoctral Research Associate
Dept. Biochemistry and Molecular Biophysics
Washington University School of Medicine
660 S. Euclid, Campus Box 8231
St. Louis, MO 63110

Re: [ccp4bb] covalent bond

[Andres Libreros]

Thanks Eleanor, John and Nigel for your tips. Actually it seems to be a problem 
of geometry restraints. 
Nigel, let me edit the .def file as John recommended and then I'll contact you. 

Best regards

Gerardo Andrés Libreros 
Laboratório de Biologia Estrutural Aplicada
Universidade de São Paulo
São Paulo- Brasil. 



Enviado desde mi iPhone

> El 7 dic 2017, a las 17:08, Nigel Moriarty  escribió:
> 
> Gerado
> 
> John is correct that you can modify the restraints model to include a 
> covalent bond. Also in more recent version of Phenix, if the input geometry 
> is approximately correct, the bond can be automatically linked.
> 
> If you send me the input file, I'll take a closer look. 
> 
> Cheers
> 
> Nigel
> 
> ---
> Nigel W. Moriarty
> Building 33R0349, Molecular Biophysics and Integrated Bioimaging
> Lawrence Berkeley National Laboratory
> Berkeley, CA 94720-8235
> Phone : 510-486-5709 Email : nwmoria...@lbl.gov
> Fax   : 510-486-5909   Web  : CCI.LBL.gov
> 
>> On Thu, Dec 7, 2017 at 9:51 AM, Tanner, John J.  
>> wrote:
>> You can add a covalent bond by modifying this part of the PHENIX .def file 
>> via the gui:
>> 
>>   geometry_restraints.edits {
>> excessive_bond_distance_limit = 10
>> bond {
>>   action = *add delete change
>>   atom_selection_1 = None
>>   atom_selection_2 = None
>>   symmetry_operation = None
>>   distance_ideal = None
>>   sigma = None
>>   slack = None
>> }
>> 
>> John J. Tanner
>> Professor of Biochemistry and Chemistry
>> Chair, Biochemistry Department Graduate Admissions Committee 
>> Department of Biochemistry
>> University of Missouri-Columbia
>> 117 Schweitzer Hall
>> 503 S College Avenue
>> Columbia, MO 65211
>> Phone: 573-884-1280
>> Fax: 573-882-5635
>> Email: tanne...@missouri.edu
>> http://faculty.missouri.edu/~tannerjj/tannergroup/tanner.html
>> Lab: Schlundt Annex rooms 3,6,9, 203B, 203C
>> Office: Schlundt Annex 203A
>> 
>>> On Dec 7, 2017, at 11:40 AM, Eleanor Dodson 
>>> <176a9d5ebad7-dmarc-requ...@jiscmail.ac.uk> wrote:
>>> 
>>> Is that format correct?
>>> Eleanor
>>> 
 On 7 December 2017 at 17:32, gerardo andres 
 <130afa955101-dmarc-requ...@jiscmail.ac.uk> wrote:
 Hi everyone. I´m trying to make a covalent bond between a cystein residue 
 of a protein and its ligand, but always that I do the refinement by 
 Phenix, the ligand is placed outside of its electronic density. I edited 
 the PDB file like this: 
 LINK SG  CYS A 215 C02 LIG C   1 , but this is not 
 working.  Someone has some idea to address this problem?
 
 Thanks,
 
 Gerardo
>>> 
>> 
> 

Re: [ccp4bb] ITC data clarification.

[Wim Burmeister]

Hello,
only a test of the biological function of mutants will tell whether  
your interface is an artefact or not.
It is very well possible that an alanine mutations increases binding;  
you have to inspect the interface carefully whether there were for  
example buried hydrogen donors or flexible residues in the WT  
interface which made the interaction less than optimal.

Regards
Wim

Dharma  a écrit :


Hello CCP4 users,
Based on the crystal structure of a two molecule protein complex, I  
have mutated (alanine substitutions) one of the putative binding  
interface. The mutant binds with much higher affinity than the wild  
type.
However, the signature plot of ITC data reveals a decrease in the  
enthalpy but increase on the entropy (deltaS). Thus overall increase  
in deltaG.

I want to know if it’s relevant biological interface or a crystal artifact.
Suggestions please.

Thanks
Regards
Dharma
Sent from my iPhone