Dear Patrick,
I did use microseeding but for a different ligand. Will try with this as
well.
Thank you
REgards
Kavya
On 2023-02-07 16:20, Patrick Shaw Stewart wrote:
>> As to why previously in a very similar condition you did get your desired
>> protein plus (other) ligand
>
> Kavya, d
>
> As to why previously in a very similar condition you did get your desired
> protein plus (other) ligand
Kavya, did you use microseeding? That's the way to get consistent results.
Since you're changing the ligand I suggest you go back and run a few random
screens (with crushed crystals of yo
gt; FROM: CCP4 bulletin board on behalf of kavyashreem
>
> DATE: Friday, February 3, 2023 at 2:32 AM
> TO: CCP4BB@JISCMAIL.AC.UK
> SUBJECT: [ccp4bb] Regarding the diffraction image
>
> Dear all,
>
> We crystallized a protein (30kDa) + ligand (by cocrystallization), in
lto:kavyashr...@instem.res.in>>
Date: Friday, February 3, 2023 at 2:32 AM
To: CCP4BB@JISCMAIL.AC.UK <mailto:CCP4BB@JISCMAIL.AC.UK> mailto:CCP4BB@JISCMAIL.AC.UK>>
Subject: [ccp4bb] Regarding the diffraction image
Dear all,
We crystallized a protein (30kDa) + ligand (by cocrystall
> SENT: Saturday, February 4, 2023 14:32
> TO: CCP4BB@JISCMAIL.AC.UK
> SUBJECT: Re: [ccp4bb] [EXT] Re: [ccp4bb] Regarding the diffraction image
>
> PS it's probably not a salt crystal...TCEP is not a salt, your ligand I
> presume is also not a salt, a small cleaved
assisted by anomalous P
(or Zn?) signal...
Have fun, BR
From: CCP4 bulletin board On Behalf Of Mark J. van Raaij
Sent: Saturday, February 4, 2023 14:32
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] [EXT] Re: [ccp4bb] Regarding the diffraction image
PS it’s probably not a salt crystal…TCEP
Dear Mark,
Thanks for the clarification.
Regards
Kavya
On 2023-02-05 04:02, Mark J. van Raaij wrote:
> PS it's probably not a salt crystal...TCEP is not a salt, your ligand I
> presume is also not a salt, a small cleaved peptide neither. As to why
> previously in a very similar conditi
PS it’s probably not a salt crystal…TCEP is not a salt, your ligand I presume
is also not a salt, a small cleaved peptide neither. As to why previously in a
very similar condition you did get your desired protein plus (other) ligand
crystal, it just means the molecule (TCEP') crystallises in a s
the unit cell in the c direction is quite long, 49 Å, this gives the relatively
close spots in one direction.
Mark J van Raaij
Dpto de Estructura de Macromoleculas, lab 20B
Centro Nacional de Biotecnologia - CSIC
calle Darwin 3
E-28049 Madrid, Spain
tel. +34 91 585 4616 (internal 432092)
Section
Dear all,
Sorry for the confusion created, I did not mean that a protein would
have fit in the small unit cell. My question was -
1. Why are there closely spaced spots arising in salt crystal?
2. If TCEP could crystallize in the condition, I have got a protein
(same as this)+ligand (differen
Dear Mark,
I did think it was salt, so I checked the same batch of protein which
went for crystallization by running a gel, it was intact, no cleavage.
My doubts arose because of two things
1. I crystallized the same protein with another ligand, in very similar
condition (10% PEG3350, 50mM Zn
Hi Jessica,
I am quite sure the protein cannot be fit in this unitcell. I was just
curious why the diffraction has closely spaced spots.
Thanks
On 2023-02-04 00:02, Jessica Bruhn wrote:
> Hi Kavya,
>
> As others have mentioned, the unit cell is too small to contain your protein.
> With
93230 Romainville
> France
> T: +33 7 81 26 95 70
> www.glpg.com [3]
>
> FROM: CCP4 bulletin board ON BEHALF OF mesters@biochem
> SENT: vendredi 3 février 2023 9:53 AM
> TO: CCP4BB@JISCMAIL.AC.UK
> SUBJECT: Re: [ccp4bb] Regarding the diffraction image
>
> Yo
Hi Kavya,
As others have mentioned, the unit cell is too small to contain your
protein. With a volume of ~4820 Ang^3, the unit cell can contain at most
~268 atoms, excluding hydrogens (divide the volume by 18 to get this
number). If the symmetry is P3, then the asymmetric unit can only contain
~89
With 50 mM Zn++ in solution, whatever it is will probably have a Zn salt in
it. So if you wanted to solve it by direct methods or via SAD - that should
do well. Sadly (hur hur) it's probably quite small, whatever it is.
Artem
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On Fri, Feb 3, 2023 at 6:20 AM
like others mentioned, looks like something in between a salt and a protein,
perhaps TCEP, the ligand, a peptide cleaved from your protein by trace protease.
If possible, I would move the detector closer, collect an atomic resolution
dataset and try to solve the structure by direct methods. You n
M
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Regarding the diffraction image
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A and B unit cell dimensions
Hi Kavya,
Try https://csb.wfu.edu/tools/vmcalc/vm.html
This tells you that a 30kD protein simply does not fit the cell.
I am pretty sure you crystallised the ligand, or TCEP actually.
Also, if you look at the diffractions pattern, its clear the crystal diffracts
beyond 1.0A, diffraction spots
A and B unit cell dimensions are hardly bigger than twice the uni cell of cubic
NaCl and will probably not accommodate a 30 kDa protein.
Make a SDS gel of washed & dissolved crystals to be sure these are not alt or
inhibitor crystals. You can stain the crystals with IzIt...
J.
--
Dr. math. et
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