Human carbonic anhdyrase II can be easily crystallized from 1.3 M sodium
citrate/0.1 M TrisCl pH 8.5 at 10 mg/mL protein concentration. Crystals
are P21 and easily diffract to beyond 2.0 A on a home source. We
cryopreserve in ML + 30% glucose. Sulfonamide ligands are easy to soak
into the crystals in a few minutes during cryopreservation, and provide
teaching opportunities for protein-ligand model building and refinement.
Cheers,
___________________________________
Roger Rowlett
Gordon & Dorothy Kline Professor
Department of Chemistry
Colgate University
13 Oak Drive
Hamilton, NY 13346
tel: (315)-228-7245
ofc: (315)-228-7395
fax: (315)-228-7935
email: rrowl...@colgate.edu
On 02/04/2013 12:31 PM, Jim Pflugrath wrote:
A number of protein crystal recipes are available on the Rigaku web site:
http://www.rigaku.com/products/protein/recipes
Lysozyme is nice because it is so cheap, grows quickly, cryoprotectants in a
straightforward way, and the unit cells are not large nor are the crystals
fragile. One can get triclinic, monoclinic, orthorhombic, and tetragonal
crystals relatively quickly.
With hen egg white lysozyme if you are not up for sulfur-SAD phasing, then
co-crystallizing or quick-soaking in iodide (say 100 mM) gives a very nice
anomalous signal at just about any wavelength. That is, no need to worry about
going to a so-called edge.
While there are many other easy-to-go crystals, I have found that none of them
combine all the properties of hen egg white lysozyme has a good teaching tool.
Jim
==========
Can you all suggest some protein crystallization conditions (along with
cryo conditions) for some commercially available proteins? I'm looking
to get 6-8 different ones (and we'll just take them and see how it
goes). I wouldn't mind knowing unit cell parameters as well (just a
citation works, I can have them figure it out). I have about 7 weeks to
get everything grown and frozen and ready to go.
Any help would be greatly appreciated. It always amazes me how helpful
this group is. Thank you very much.
Dave
