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 <[EMAIL PROTECTED]>
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: [EMAIL PROTECTED]
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)
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