Right on target. Many proteins require some ionic strength in the
solution to maintain solubility and prevent protein aggregation. Usually
NaCl is used for this purpose. You can also use glycerol to enhance
solubility, but this may interfere with crystallization if that's the
next step. NaCl usually interferes less with crystallization. The
minimum concentration of salt required will have to be determined
empirically.
For gel exclusion chromatography, it is usually necessary to include at
least 50 mM NaCl in the buffer to prevent non-specific adsorption to the
gel medium. I use 100 mM in my GEC columns. If your forget to add the
salt, your elution times will look very wonky, especially if you are
trying to use elution data to estimate molecular weight.
Cheers,
_______________________________________
Roger S. 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: [email protected]
On 11/20/2013 4:37 PM, Tim Gruene wrote:
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Hello Dhana,
use a buffer at least, and some salt - if I remember correctly, gel
filtration resins require 150mM salt concentration for proper
separation (might have been 50mM). Even 150mM may be too little to
keep your protein soluble.
Best,
Tim
On 11/20/2013 09:56 PM, Dhanasekaran Varudharasu wrote:
Dear Crystallographers,
I dialysed a 30 kDa protein (Recombinant protein which was eluted
by 20 mM Tris, 500 mM NaCl, 120 mM imidazole) against water for
overnight. But it gets precipitated after 12 hours. Can anybody
give some suggestion to avoid precipitation. Thanks Dhana
- --
Dr Tim Gruene
Institut fuer anorganische Chemie
Tammannstr. 4
D-37077 Goettingen
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