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[ccp4bb]: SUMMARY (II) Hg compound
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Sorry, I clearly misunderstood what Louis Renault proposed about "soaking
prior to crystallisation". He has kindly sent me a thorough explanation:
> 1- to expose at 4 degrees for few hours your protein with a very small
> excess of heavy-atoms in the normal protein stock solution
> (it means you just add to your protein your heavy-atom solubilized not in the mother
> liquor otherwise you induce crystallization, but in the same
> buffer that your normal protein stock solution. You do this
> to have a small ratio of (protein/heavy-atoms) from 1/5 to 1/1. )
> 2- to crystallize afterwards this putative complexe (protein+Hg) in the
> usual mother liquor.
> The advantage of this method is that there is no other competitor
> at this early step from the mother liquor to chelate heavy-atoms.
> Therefore heavy-atoms bind only to the protein in theory
> and a small ratio of (protein/heavy-atoms) is required, whereas when
> you crytallize with heavy-atom in your mother liquor, heavy-atom can
> be chelated by citrate, PO4, tris, ... if these coumpound are present
> in the mother liquor and you would require higher concentration.
> There is a very small paragraph about this method in the
> CCP4 Daresbury Study Weekend Proceedings from 1991
> Isomorphous replacement and anomalous scattering. DL/SCI/R32
> in one article about preparation of heavy-atom solutions
> (Sorry I do not have the book with me to tell which of the article
> is the proper reference).
> You can also crystallize your protein+mother liquor+heavy-atom in mother liquor
> as you told, but here you need usually higher heavy-atom concentration
> because of the mother liquor which contains often some chelating agent as well.
> good luck
Thank you very much Louis!
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