Matt,
To add to George's advice, this post concerns ligand exchange done in
cryosolutions:
1. The first step is to find cryosolutions in which your crystals are
stable for 20 mins or longer.
2. Solubilize the ligand you want to exchange in the cryosolution and
verify that it is soluble (1mM concentrations will ensure that it will get
in).
3. Place the crystal in the solution for 5 min. Here you are starting to
wash out the old ligand and exchange it for the new one.
4. Move the crystal into a new clean cryo-ligand solution. (5 min)
5. More again to a clean one. (use a longer soak if you so please)
To verify that the procedure works, or to improve it, it would be nice
that one of the two ligands to has an anomalous scatterer. Bromine or
iodine would be useful.
The publications below will give you further details regarding the
procedure to follow:
148. Ciccone L., Tepshi L., Nencetti, S. & Stura E.A. (2015) Transthyretin
complexes with curcumin and bromo-estradiol: Evaluation of solubilizing
multicomponent mixtures New Biotech. 32:54—64.
http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=25224922
http://dx.doi.org/10.1016/j.nbt.2014.09.002
149 Ciccone L., Vera L., Tepshi L., Rosalia L., Rossello A. & Stura E.A.
(2015) Multicomponent mixtures for cryoprotection and ligand
solubilization. Biotechnology Reports 7:120—127.
http://dx.doi.org/10.1016/j.btre.2015.05.008
Hope this is useful,
Enrico.
On Wed, 16 Mar 2016 19:37:57 +0100, Matthew Bratkowski
<[log in to unmask]> wrote:
> Hello all,
>
> I am looking for information about soaking a competitor ligand into
> protein
> crystals that contain a different ligand that is presumably at the same
> site, allowing for exchange of the ligands. I was wondering if anyone
> could suggest a reference that contains a good general procedure for this
> technique or could provide some personal experience as to how successful
> this technique is.
>
> Thanks,
> Matt
--
Enrico A. Stura D.Phil. (Oxon) , Tel: 33 (0)1 69 08 4302 Office
Room 19, Bat.152, Tel: 33 (0)1 69 08 9449 Lab
LTMB, SIMOPRO, IBiTec-S, CE Saclay, 91191 Gif-sur-Yvette, FRANCE
e-mail: [log in to unmask] Fax: 33 (0)1 69 08 90 71
Proxima-2A, Soleil Synchrotron. Tel: 33 (0)1 69 35 8180 Beamline
http://scholar.google.com/citations?hl=en&user=Kvm06WIoPAsC&pagesize=100&sortby=pubdate
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