Re: [ccp4bb] Density around PEG

[Prem Prakash]

What is your occupancy of PEG here ?,you should also look at it.

Best
Prem

On Thu, Dec 14, 2017 at 6:37 AM, sanjeev kumar 
wrote:

> Thanks, I'll check it out.
>
> On Wed, Dec 13, 2017 at 8:06 PM Anthony Addlagatta <
> anthonyaddlaga...@gmail.com> wrote:
>
>> This could be 2-mercaptoethanol.
>>
>> Anthony
>>
>> On Wed, Dec 13, 2017 at 11:24 PM, sanjeev kumar 
>> wrote:
>>
>>> Hi all,
>>> Can someone suggest what could be the green density around PEG?
>>> Protein was purified in Tris, Nacl & 2-Mercaptoethanol and Crystallization
>>> condition contains PEG, Magnesium acetate and Potassium Iodide. Image are
>>> attached below.
>>> Thanks
>>>
>>> best
>>> Sanjeev Kumar, PhD
>>>
>>>
>>
>>
>> --
>> Dr. Anthony Addlagatta
>> Principal Scientist
>> Center for Chemical Biology
>> CSIR-Indian Institute of Chemical Technology [IICT]
>> Tarnaka, Hyderabad
>> Telangana-500 607, INDIA
>> Tel:91-40-27191860
>> Web: http://www.iictindia.org/staffprofiles/staffprofile.
>> aspx?emp_id=IICT1735
>>  https://sites.google.com/site/chembioliict/home/dr-anthony-
>> addlagatta-1
>>
> --
> Sanjeev Kumar, PhD
> C/o Prof. S. Gourinath
> Lab No # 430
> School of Life Sciences
> JNU, New Delhi-110067
> Contact No-+919650747960
>

Re: [ccp4bb] coordinate transformation

[Paul Emsley]

On 13/12/2017 13:50, Kajander, Tommi A wrote:

Hello,

If someone could point this out would be very helpful... Wasnt there a simple script somewhere that would 
transfer coordinates close to origin - if they for some reason are not? Just cant find anything right away. 



At the risk of not answering the question because it's not a simple script, my 
I recommend Coot?

File -> Open -> yourcoords.cif
Draw -> Cell & Symm -> Master Switch -> Yes
Show Unit Cells -> Yes
OK
Drag the View to the Origin # it's marked with an "O"
Middle-mouse click on an Symmetry-related Atom # that's close to the origin
Extensions -> Modelling -> Symm Shift Reference Chain Here

Re: [ccp4bb] Density around PEG

[sanjeev kumar]

Thanks, I'll check it out.

On Wed, Dec 13, 2017 at 8:06 PM Anthony Addlagatta <
anthonyaddlaga...@gmail.com> wrote:

> This could be 2-mercaptoethanol.
>
> Anthony
>
> On Wed, Dec 13, 2017 at 11:24 PM, sanjeev kumar 
> wrote:
>
>> Hi all,
>> Can someone suggest what could be the green density around PEG?   Protein
>> was purified in Tris, Nacl & 2-Mercaptoethanol and Crystallization
>> condition contains PEG, Magnesium acetate and Potassium Iodide. Image are
>> attached below.
>> Thanks
>>
>> best
>> Sanjeev Kumar, PhD
>>
>>
>
>
> --
> Dr. Anthony Addlagatta
> Principal Scientist
> Center for Chemical Biology
> CSIR-Indian Institute of Chemical Technology [IICT]
> Tarnaka, Hyderabad
> Telangana-500 607, INDIA
> Tel:91-40-27191860
> Web:
> http://www.iictindia.org/staffprofiles/staffprofile.aspx?emp_id=IICT1735
>
> https://sites.google.com/site/chembioliict/home/dr-anthony-addlagatta-1
>
-- 
Sanjeev Kumar, PhD
C/o Prof. S. Gourinath
Lab No # 430
School of Life Sciences
JNU, New Delhi-110067
Contact No-+919650747960

Re: [ccp4bb] Overlapping ligand electron density

[Diana Tomchick]

Matt,

Modeling two molecules that occupy overlapping binding sites in a structure 
simply involves designating them as alternate conformers, with the same chain 
and residue number, and an occupancy that sums to 1.0. For example, if you have 
an AMP and an ADP that occupy the same binding site, you would define them as

AAMP B 501
BADP B 501

and initially set the occupancies for the atoms in each conformer to the ratio 
(50:50, 30:70, etc.) that you observe in the density.

Refinement in this manner is straightforward in PHENIX.

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 13, 2017, at 1:11 PM, Matthew Bratkowski 
> wrote:

Hello all,

I am working on a ligand binds near the active site of the protein, such that 
part of the ligand would clash with part of the natural substrate.  I recently 
co-crystallized the enzyme with both molecules and solved the crystal structure 
to high resolution (around 1.4 angstrom).  Surprisingly, the structure appears 
to contain both molecules.  A few atoms from both molecules are located only 
~1.4 A apart and are clashing (although not overlapping).  The electron density 
between them looks connected, but based on the two groups that are clashing (a 
methyl group and a carbonyl oxygen), I do not think that a covalent adduct 
occurs.  I had a few questions.

1) My guess is that the crystal is "sampling" two different conformational 
states and that both are visible due to the high diffraction resolution.  The 
substrate contains a ring that shows a characteristic "hole" in the electron 
density and binds in the exact substrate binding site, suggesting that it is 
not a different molecule (no molecules with ring structures were included in 
the sample, crystallization buffer, or cry-protectant).  One of the two 
proteins in the ASU contains electron density for whole substrate, while the 
other site has only density around the ring.  However, a sizable amount of red 
FoFc density is present around the substrate, suggesting that it is only 
partially occupied.

Does this explanation seem plausible?

2) How would I go about modeling these two molecules in the structure?  Should 
I include both molecules (in their entirety) in the structure?  I suspect that 
neither the ligand nor substrate are completely occupied, so should I modify 
the occupancies to reflect this?

Thanks,
Matt




UT Southwestern


Medical Center



The future of medicine, today.

Re: [ccp4bb] Overlapping ligand electron density

[Bernhard Rupp]

Only two comments:

 

a.  At that resolution, constrained group occupancy refinement should work 
reasonably well (provided you can model the 2 entities). Then you also do not 
have clashes between the molecules, because Occ(A)+Occ(B)=1, meaning when one 
(A) is there, the other one (B) is not. This works with refmac (external 
keyword file); if you need more sophisticated occupancy re/constraints SHELXL 
may offer more opportunities.
b.  There is no necessity for the two NCS copies of the binding site to 
look exactly the same (non-equivalent). Maybe there is a good reason/story 
(accessibility, contacts etc) for one site to be occupied differently than the 
other one. 

 

Best, BR

 

From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Matthew 
Bratkowski
Sent: Wednesday, December 13, 2017 11:12 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] Overlapping ligand electron density

 

Hello all,

 

I am working on a ligand binds near the active site of the protein, such that 
part of the ligand would clash with part of the natural substrate.  I recently 
co-crystallized the enzyme with both molecules and solved the crystal structure 
to high resolution (around 1.4 angstrom).  Surprisingly, the structure appears 
to contain both molecules.  A few atoms from both molecules are located only 
~1.4 A apart and are clashing (although not overlapping).  The electron density 
between them looks connected, but based on the two groups that are clashing (a 
methyl group and a carbonyl oxygen), I do not think that a covalent adduct 
occurs.  I had a few questions.

 

1) My guess is that the crystal is "sampling" two different conformational 
states and that both are visible due to the high diffraction resolution.  The 
substrate contains a ring that shows a characteristic "hole" in the electron 
density and binds in the exact substrate binding site, suggesting that it is 
not a different molecule (no molecules with ring structures were included in 
the sample, crystallization buffer, or cry-protectant).  One of the two 
proteins in the ASU contains electron density for whole substrate, while the 
other site has only density around the ring.  However, a sizable amount of red 
FoFc density is present around the substrate, suggesting that it is only 
partially occupied.

 

Does this explanation seem plausible?

 

2) How would I go about modeling these two molecules in the structure?  Should 
I include both molecules (in their entirety) in the structure?  I suspect that 
neither the ligand nor substrate are completely occupied, so should I modify 
the occupancies to reflect this?

 

Thanks,

Matt   

Re: [ccp4bb] Overlapping ligand electron density

[Pearce, N.M. (Nick)]

It’s definitely possible to have a superposition of states: 
https://www.ncbi.nlm.nih.gov/m/pubmed/28291761/

You need to use alternate conformers to generate the different states of the 
crystal.

Thanks,
Nick

On 13 Dec 2017, at 20:12, Matthew Bratkowski 
> wrote:

Hello all,

I am working on a ligand binds near the active site of the protein, such that 
part of the ligand would clash with part of the natural substrate.  I recently 
co-crystallized the enzyme with both molecules and solved the crystal structure 
to high resolution (around 1.4 angstrom).  Surprisingly, the structure appears 
to contain both molecules.  A few atoms from both molecules are located only 
~1.4 A apart and are clashing (although not overlapping).  The electron density 
between them looks connected, but based on the two groups that are clashing (a 
methyl group and a carbonyl oxygen), I do not think that a covalent adduct 
occurs.  I had a few questions.

1) My guess is that the crystal is "sampling" two different conformational 
states and that both are visible due to the high diffraction resolution.  The 
substrate contains a ring that shows a characteristic "hole" in the electron 
density and binds in the exact substrate binding site, suggesting that it is 
not a different molecule (no molecules with ring structures were included in 
the sample, crystallization buffer, or cry-protectant).  One of the two 
proteins in the ASU contains electron density for whole substrate, while the 
other site has only density around the ring.  However, a sizable amount of red 
FoFc density is present around the substrate, suggesting that it is only 
partially occupied.

Does this explanation seem plausible?

2) How would I go about modeling these two molecules in the structure?  Should 
I include both molecules (in their entirety) in the structure?  I suspect that 
neither the ligand nor substrate are completely occupied, so should I modify 
the occupancies to reflect this?

Thanks,
Matt

[ccp4bb] Overlapping ligand electron density

[Matthew Bratkowski]

Hello all,

I am working on a ligand binds near the active site of the protein, such
that part of the ligand would clash with part of the natural substrate.  I
recently co-crystallized the enzyme with both molecules and solved the
crystal structure to high resolution (around 1.4 angstrom).  Surprisingly,
the structure appears to contain both molecules.  A few atoms from both
molecules are located only ~1.4 A apart and are clashing (although not
overlapping).  The electron density between them looks connected, but based
on the two groups that are clashing (a methyl group and a carbonyl oxygen),
I do not think that a covalent adduct occurs.  I had a few questions.

1) My guess is that the crystal is "sampling" two different conformational
states and that both are visible due to the high diffraction resolution.
The substrate contains a ring that shows a characteristic "hole" in the
electron density and binds in the exact substrate binding site, suggesting
that it is not a different molecule (no molecules with ring structures were
included in the sample, crystallization buffer, or cry-protectant).  One of
the two proteins in the ASU contains electron density for whole substrate,
while the other site has only density around the ring.  However, a sizable
amount of red FoFc density is present around the substrate, suggesting that
it is only partially occupied.

Does this explanation seem plausible?

2) How would I go about modeling these two molecules in the structure?
Should I include both molecules (in their entirety) in the structure?  I
suspect that neither the ligand nor substrate are completely occupied, so
should I modify the occupancies to reflect this?

Thanks,
Matt

Re: [ccp4bb] coordinate transformation

[Bartosz Sekula]

Hi Tommy,
Try ACHESYM, it is very useful:
http://achesym.ibch.poznan.pl/

Here is the link to the publication:
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4257622/

Good luck,
Bartosz

Re: [ccp4bb] coordinate transformation

[Keller, Jacob]

Wouldn't the PISA server do something like this?

JPK

From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Kajander, 
Tommi A
Sent: Wednesday, December 13, 2017 8:50 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] coordinate transformation


Hello,

If someone could point this out would be very helpful... Wasnt there a simple 
script somewhere that would transfer coordinates close to origin - if they for 
some reason are not? Just cant find anything right away. Sure i have done this 
before...



Thanks,

Tommi

[ccp4bb] coordinate transformation

[Kajander, Tommi A]

Hello,

If someone could point this out would be very helpful... Wasnt there a simple 
script somewhere that would transfer coordinates close to origin - if they for 
some reason are not? Just cant find anything right away. Sure i have done this 
before...


Thanks,

Tommi

Re: [ccp4bb] PyMol question

[Johannes Cramer]

you can try the 'hide cell' command. This should hide the unit cell box,
however, I don't know why it is not displayed in your 'unrendered' gui...

Cheers,
Johannes

2017-12-11 23:47 GMT+01:00 Cygler, Miroslaw :

> Hi,
> When I loaded the ed map into PyMol v1.8.4.1 and used iso mesh around part
> of the protein all is well until I use the RAY command. In the ray traced
> image I see the unit cell box that does not show on the image in the normal
> view. How can I remove the box from the ray traced image?
> Thanks for your help,
>
> Mirek
>
>
>
>