His-tagged proteins....

Frank Fuerst ffrank at rz.uni-potsdam.de
Fri Mar 23 07:20:40 EST 2001


[quoting adjusted, please don't produce TOFU (1)]

Michael Witty <mw132 at mole.bio.cam.ac.uk> wrote:

>On 22 Mar 2001, Debashis Mukhopadhyay wrote:
>
[...]
>> I have a protein domain which is cloned and expressed
>> in E.coli having an N-terminal His-tag and a Thrombin
>> cleavage site just after that. The idea is to get rid
>> of the tag for crystallisation purposes by thrombin
>> treatment. Can you point me to any reference(s), ideal
>> for a novice, having details for the same and
>> subsequent purification steps to eliminate the
>> contaminants! Any nice protocol would as well be good!

>. . . a lot of people would say, don't bother removing the tag if you can
>demonstrate activity with the tag present.  Can you easily do that?  Mike.


But the hist tag might well interfere with crystallization (it might
even improve it...).

Bye, Frank


(1) TOFU: no, not the meat replacement, it's German usenet jargon for
"Text oben, Fullquote unten" (Text above, Fullquote below, or perhaps
'Text over Fullquote underneath')




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