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glutathione fusion protein

Tony Hodge tph at mrc-lmb.cam.ac.uk
Fri Jul 8 04:03:16 EST 1994


Subject: glutathione fusion protein
From: D.R. Bell, mbdxb at s-crim1.dl.ac.uk
Date: 7 Jul 1994 13:15:54 GMT
In article <2vgv6a$fdq at mserv1.dl.ac.uk> D.R. Bell,
mbdxb at s-crim1.dl.ac.uk writes:
>We have a GST fusion protein which comes out as insoluble.
>
>Has anyone tried solubilising such proteins, and then trying
affinity
>purification of the refolded GST-fusion protein?
>
>Any comments and protocols welcome!


Yes we have tried this and we too have big problems.  So far two
different constructs made by two people have failed to be purified
on affinity purification columns.  In my case I dialysed down to 2M
Urea and my colleague to 500mM from an initial 8M and were unable to
show binding to the column.  Pharmacia have been unable to give any
help when contacted them.  We have had success with one soluble
fusion construct where the column behaved as expected.

If anyone can advise on how to get this working we would be most
grateful to hear from them.




.
Tony P Hodge
Structural Studies Division
Medical Research Council Laboratory of Molecular Biology
Hills Road
Cambridge
CB2  2QH
UK

Phone (0223)  402260

Fax     (0223)  213556



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