Structure of His-Tag complex

Cornelius Krasel krasel at wpxx02.toxi.uni-wuerzburg.de
Thu May 8 10:28:50 EST 1997


Thorsten Schmidt (Thorsten.Schmidt at rz.ruhr-uni-bochum.de) wrote:
> When one loads the column with a protein mixture only the target protein
> forms with its histidines a complex with the nickel and the resin.
> After washing steps the protein can be eluted with a buffer containing
> Imidazole.
> 
> I want to know the exact structure of this complex and the reason, why
> it is possible to elute the protein with Imidazole.

Depending on the resin that you use, the Ni2+ is complexed by two (IDA),
three (NTA) or four (TALON) valences; that is, there are four (IDA),
three (NTA) or two (TALON) left for the histidines to bind. The
chemical structure of the complex can be found in the various leaflets
given away by the companies who want to sell that stuff. If you mean
"structure" in terms of 3 D coordinates, I have been told that "His-Tags"
are usually pretty flexible and therefore do not show up in X-ray
structures.

The reason why it is possible to elute the protein with imidazol is
pretty obvious if you have a look at the chemical structure of it.
It competes with the histidine for the metal ion.

--Cornelius.

-- 
/* Cornelius Krasel, U Wuerzburg, Dept. of Pharmacology, Versbacher Str. 9 */
/* D-97078 Wuerzburg, Germany   email: phak004 at rzbox.uni-wuerzburg.de  SP3 */
/* "Science is the game we play with God to find out what His rules are."  */



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