Trombin cleavage of fusion proteins

Song Tan tan at aeolus.vmsmail.ethz.ch
Fri Sep 2 04:35:38 EST 1994


In article <9409012138.AA65559 at acs4.acs.ucalgary.ca>,
msseyed at ACS.UCALGARY.CA ("Soheil Seyed Mahmoud") wrote:
                    
> Hi there. I have expressed a protein (ca 22kd) as a fusion with
> GST in E.coli. I can purify ther fusion protein on agarose bound
> glutathione beads. However, as I attempt to add the thrombin
> bufer (20mMtris (pH8.3), 5mM CaCl2, with or without 150mMNacl),
> the fusion protein precipitates out. Any suggestions how I can
> get over this problem? Thanmks a bunch.

How about cleaving the fusion protein while it's still bound to the
glutathione beads?  Pharmacia's "GST Gene Fusion System" booklet on the
pGEX expression system describes the procedure for doing this.

-- 
Song Tan
Institute for Molecular Biology and Biophysics
ETH-Honggerberg (Swiss Federal Institute of Technology)
8093 Zurich, Switzerland
email:  tan at aeolus.vmsmail.ethz.ch



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