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Jerry McCully wrote:
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Dear ccp4bb experts:
I'm crystallizing a dynamic protein-protein complex. The two
components bind to each other
with a micro-molar affinity(This affinity was shown by Biacore3000
analysis, but I can not purify the complex by gel-filtration).
I can get their crystals, respectively. But so far I have not
got any crystals of the complex even if I tried several molar ratios
between them.
When I used the complex sample for crystallization, the smaller
component was easily grown from the conditions containing PEG3350, and
the other from MPD.
Does anyone have the experience to crystallize this kind of
low-affinity protein complexes?
Any suggestion will be highly appreciated.
Jerry McCully
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Dear Jerry
usually the high protein concentrations encountered under
crystallization conditions will always drive the system to complex
formation (even for instable complexes with high dissociation
constants). Obviously, as well as some proteins do not crystallize some
complexes won't do so either. It should be sufficient to screen a high
number of crystallization conditions, usually the conditions of complex
crystallisation will be very different from the conditions of
crystallization of the individual compounds. I would be a bit worried
about the Biacore result, if it is a low micromolar dissociation
constant, I would expect that it should be possible to observe the
complex by gel filtration. It would be a good idea to try another method
for complex characterization (EM, microcalorimetry etc.)
Yours
Wim Burmeister
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Wim Burmeister
Professeur, Membre de l'Institut Universitaire de France
Laboratoire de Virologie Moleculaire et Structurale FRE 2854 CNRS-UJF
c/o EMBL, 6 rue Jules Horowitz
B.P. 181, F-38042 Grenoble Cedex 9 FRANCE
E-mail: [EMAIL PROTECTED]
Tel: +33 (0) 476 20 72 82 Fax: +33 (0) 476 20 71 99
http://www2.ujf-grenoble.fr/pharmacie/laboratoires/gdrviro
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