Hi Raphael
I can second Tom and Matthews comments. We got crystals only after including an
ion exchange column (even though we work with a GST construct cleaved off with
thrombin). Well diffracting crystals then only grow by seeding in reduced
precipitant concentration. Additionally, the seeds allow us to test for
additive salts in which initial crystals did not pop up. I often use a whole
set of fine screens with different salts in which I am seeding in the protein.
And in almost all cases, the seeding yields in good crystals only after also
changing the protein concentration (say, from initially 12mg/ml to 8-30mg/ml
with reduced precipitant concentration). You did not mention what you changed
in your seeding experiment, but try a bunch of different conditions,
concentrations (and temperature) with the seeds.
Matthias
Dr. Matthias Barone
AG Kuehne, Rational Drug Design
Leibniz-Forschungsinstitut für Molekulare Pharmakologie (FMP)
Robert-Rössle-Strasse 10
13125 Berlin
Germany
Phone: +49 (0)30 94793-284
________________________________
From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> on behalf of hoh
<h...@cbs.cnrs.fr>
Sent: Friday, December 18, 2020 9:46:58 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Micro/Macro crystal seeding experience
Hi Raphael
Think about simple refinement,
differents kits are available
additifs kits:
- additif, Gras and Silver bullet : from Hampton research
- Angstrom and Morpheus from MDL
- JBScreen Plus from Jena bioscience
detergent kits:
- from Hampton resaerch
- from Jena bioscience
FH
________________________________________
François Hoh
Centre de Biochimie Structurale,
UMR 5048 CNRS, UMR 1054 INSERM
29, rue de navacelles
34090 Montpellier Cedex, France.
Phone: +33 467 417 706
Fax: +33 467 417 913
________________________________________
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