how to cleave His-tag

Leemor Joshua-Tor leemor at CSHL.ORG
Wed Jul 17 08:19:30 EST 1996


>We'd like to engineer a cleavage site to remove the his-tag from
>our protein but cannot use a thrombin, fXa, or enterokinase site
>since these are calcium-requiring enzymes and we need to keep calcium
>from our protein.  Does anyone have a suggestion for a non-calcium
>dependent protease that we can use, and which has been successfully
>used by others (we are considering trypsin and chymotrypsin, but
>are likely to have cleavage at more than the one desired site)?
>
>Many thanks!
>Vivien yee

We used a TEV site between the his-tag and our protein. TEV
is very specific and clean. It will leave a gly on the protein.
The specificity is : glu-asn-leu-tyr-phe-gln-I-gly  (I is the cut).

In our case we ended up not having to cleave the his tag off since
we got crystals with the his tag and protease site anyway.

                                                        Leemor



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  Leemor Joshua-Tor, Ph.D.
  Cold Spring Harbor Laboratory           Tel. (516) 367 8821
  1 Bungtown Road                         Fax  (516) 367 8873
  Cold Spring Harbor, NY 11724            e-mail: leemor at cshl.org
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