Re: [ccp4bb] Help with reducing crystal mosaicity

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

Previous message:
Subject: Re: Help with reducing crystal mosaicity
From: Patrick Shaw Stewart Patrick {- at -} DOUGLAS {- dot -} CO {- dot -} UK
Date: 2007-07-11

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Subject: Re: Help with reducing crystal mosaicity
From: "Benini, Stefano" Stefano {- dot -} Benini {- at -} ASTRAZENECA {- dot -} COM
Date: 2007-07-11

Dear Mary,

the problem encountered with cryoprotectans is the change in the solution surrounding your crystal as they may not be present in your crystallisation conditions or you need to increase their concentration to make them act as cryoprotectant.
So I don't thik is the cryoprotectant itself as it is you mother liquor..

test it in a capillary so you could rule out that freezing is the problem

of course you don't want to find new crystallisation conditions when you tried everything and I know what it means!!

I would try to improve crystal quality using additives or ligand etc....,

your long axis is very worrying as well!!! may be some magic additive could change the packing and givew better diffraction too

I also had crystals growing from MPD, unfortunately there was a non-cleavable his-tag
so I could not hope that metals would help..... and other additives did not make any difference
then my contract finished and so did the project!

I hope you still have a lot of time available to try different things but I would not waste time trying to change your cryo

ciao

Stefano

Stefano Benini PhD
AstraZeneca UK
Structural Biology
Mereside 50S38
Alderley Park

-----Original Message-----
From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK]On Behalf Of
Patrick Shaw Stewart
Sent: 11 July 2007 12:10
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Help with reducing crystal mosaicity

Just a thought, Mary - going back to your original question about MPD. I extracted the crystallization conditions from REMARK 280 of 3939 PDB entries a couple of years ago. The average concentration of the MPD used was high - 38.6%, while PEGs tended to be used at lower concs, e.g. PEG400 25.7%. You can see the data at www.douglas.co.uk/top14.htm

I thought this information could be useful if you want to replace some of the MPD with another precipitant (or cryoprotectant).

Best wishes

Patrick Shaw Stewart

--
patrick@douglas.co.uk Douglas Instruments Ltd.
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Directors: Peter Baldock, Patrick Shaw Stewart, James Smith
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> -----Original Message-----
> From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Mary
> Fitzgerald
> Sent: 09 July 2007 23:05
> 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

CCP4bb navigation

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

Previous message:
Subject: Re: Help with reducing crystal mosaicity
From: Patrick Shaw Stewart Patrick {- at -} DOUGLAS {- dot -} CO {- dot -} UK
Date: 2007-07-11

Next message:
Subject: Metal-ligand(s) positioning following refinement...
From: James Pauff pauffjm {- at -} YAHOO {- dot -} COM
Date: 2007-07-11