Hi Chandan,
What do you mean by problem in crystallization? Too much precipitate, too many clear drops, ...? If you state the specific problems you are experiencing it is easier to make specific suggestions. By 388 buffers, do you mean that you have only tried 388 crystallization conditions or ~4 commercial screens? That really is not a lot of conditions to try these days. Have you tried with and without Zinc? Are there other known ligands for your protein you can try to use? Have you experimented with other variables such as protein concentration, temperature, ratio of protein solution to reservoir solution, volume of drop, crystallization method (hanging drop vs sitting drop vs. batch vs. ...), etc. Perhaps the buffer your protein is in going into the crystallization experiments needs to be optimized. Perhaps it is your protein itself that is problem. Do you have a cleavable affinity tag for purification?
Try crystallizing both the cleaved and non-cleaved protein. You may need to design other constructs of your protein. Move the tag to N- or C-terminus. Make a series of truncation mutants. Maybe you have disordered domains that are preventing crystallization - remove them. Are there homologous proteins in other species you could try? And the list goes on - there is a near limitless number of things that can be tried.
hope that will at least get you started.
good luck.
Eric
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Eric T. Larson, PhD
Biomolecular Structure Center
Department of Biochemistry
Box 357742
University of Washington
Seattle, WA 98195
email: larso...@u.washington.edu
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On Thu, 27 Jan 2011, chandan kishore wrote:
Hi Everyone,
I have to crystallize one protein which contains a zinc fingure motifs, I am
facing a problem in crystallization. I already used 388
buffers . Can anyone suggest me some books or papers freely available online on
Buffers required for crystallization.
Thanks
