His-tagged proteins

[Cornelius Krasel]

Cara Baker (crbaker at mail.tcd.ie) wrote:
> Has anyone had experience with his-tagged proteins that precipitate 
> immediately on elution with imidazole? Is there a way to get around 
> this?

Maybe elution with EDTA would work better? You can always recharge
your NTA column with NiCl2.

> Also are there any other tricks to removing backround contaminants 
> (under native conditions) apart from use of b-mercaptoethanol and 
> higher salt concs?

A Q sepharose column does the trick for me (purification of his6-
tagged Go-alpha).

(Well, I can't say that I don't have problems. I have pure protein,
 but only 5% of it is functional :-/ )

--Cornelius.

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/* Cornelius Krasel, U Wuerzburg, Dept. of Pharmacology, Versbacher Str. 9 */
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