Re: [ccp4bb] Help with reducing crystal mosaicity

- Protein crystallography

Main steps:

- Protein purification
- Crystallisation

Special:

- Programs for crystallography
- X-ray detectors

Basic tutorials:

   - Chemistry
   - Protein
   - Peptide
   - Amino Acids

Xtal community:

- CCP4BB

CCP4bb navigation

CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999

Previous message:
Subject: Post Doc on Hsp90-Complexes at ICR, London
From: Laurence Pearl Laurence {- dot -} Pearl {- at -} ICR {- dot -} AC {- dot -} UK
Date: 2007-07-10

Next message:
Subject: Re: Help with reducing crystal mosaicity
From: Edward Snell esnell {- at -} HWI {- dot -} BUFFALO {- dot -} EDU
Date: 2007-07-10

Subject: Re: Help with reducing crystal mosaicity
From: Flip Hoedemaeker flip {- at -} KEYDP {- dot -} COM
Date: 2007-07-10

You might also want to loook into using parathone for freezing... Or
collection at 260K indeed!

Flip

-----Original Message-----
From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of
mesters
Sent: Tuesday, July 10, 2007 11:01
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Help with reducing crystal mosaicity

Mary,

freezing habitually increases mosaicity. In your case, the high water
content adds to the problem.
Try not to freeze the crystal but collect at sub-zero temperature (in
short glass capillaries or use oil plugs instead).
You have to optimize the "close to freezing" data-collection temperature.

I collected complete synchrotron datasets (of GCPII in buffer with
PEG1500 and PEG400) at 260-263 Kelvin which resulted in mosaicity values
of as small as 0.07 degrees! At 277 K, the crystals only last for a few
images and freezing did not work (for the buffer mentioned before).

- J. -

> -----Original Message-----
> From: Mary Fitzgerald
> Date: Mon, 9 Jul 2007 18:05:10
> To:CCP4BB@JISCMAIL.AC.UK
> Subject: [ccp4bb] Help with reducing crystal mosaicity
>
> Help please!
>
> I'm looking for some new ideas. I have crystals that come out of a
> sitting drop with a mixture of sodium cacodylate at pH 6.5, magnesium
> acetate and MPD for the well solution. The MPD concentration is
> sufficient to act as a cryoprotectant. Currently, I directly freeze
> these crystals in liquid nitrogen. When I collect data, I typically
> have high anisotropic mosaicity; it ranges from 0.8 to 1.2. This is
> further complicated with a weakly diffracting crystal (4-5 A) that has
> a long unit cell axis of ~500 and often twinning.
>
> It has been suggested to me that the cryoprotectent is a problem. I
> haven't checked the diffraction at room temperature, yet. Please no
> suggestions of finding a different crystal form as that's not a
> consideration at the moment. I have my reasons. I did find one
> crystal that has lower mosaicity (0.5 to 0.8) but had weaker
> diffraction then the typical crystal. Attempts at flash cryoannealing
> have not helped.
>
> So, what's a good way to change the cryoprotectant if the
> cryoprotectant is the precipitant? I've considered trying dehydration
> but wasn't certain if that would help with the mosaicity.
>
> Thanks for any ideas,
>
> Mary X. Fitzgerald
> Postdoctoral Associate
>

--
Jeroen Raymundus Mesters, Ph.D.
Institut fuer Biochemie, Universitaet zu Luebeck
Zentrum fuer Medizinische Struktur und Zellbiologie
Ratzeburger Allee 160, D-23538 Luebeck
Tel: +49-451-5004070, Fax: +49-451-5004068
E-mail: mesters@biochem.uni-luebeck.de
Http://www.biochem.uni-luebeck.de
Http://www.iobcr.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 navigation

CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999

Previous message:
Subject: Post Doc on Hsp90-Complexes at ICR, London
From: Laurence Pearl Laurence {- dot -} Pearl {- at -} ICR {- dot -} AC {- dot -} UK
Date: 2007-07-10

Next message:
Subject: Re: Help with reducing crystal mosaicity
From: Edward Snell esnell {- at -} HWI {- dot -} BUFFALO {- dot -} EDU
Date: 2007-07-10