Restriction proteases?

Leemor Joshua-Tor leemor at CSHL.ORG
Sun Jan 28 12:15:18 EST 1996


>We look for the most specific way to cleave a GST fusion protein. Currently
>we use a Thrombin "restriction" site which works fine, unfortunately
>the Thrombin cleaves also somewhere inside the fusion protein. I have
>been told that the specificity of Thrombin is not too high. Therefore
>I currently wonder if I should replace the Thrombin cleavage site with
>a recognition sequence for another protease. I wonder which protease
>could others recommend? (Factor Xa seems to be common but I have heard
>that it often works not very well. What about IgA protease or entero-
>kinase?)
>
>Thanks for your help,
>
>--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."  */



There is a very nice protease, which is very specific, and so very
useful for what you want to do called TEV protease. Highly recommended.
It will not cut in any known protein and is the purest and
the best buy of units activity. You can get it from Life Technologies (BRL).
Much less can be used than recommended as you can incubate it for long
periods as there is no contaminating activity.


*******************************************************************
  Leemor Joshua-Tor
  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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