In article <37A1A4F8.29B8 at uctgsh1.uct.ac.za>, lester at uctgsh1.uct.ac.za wrote:
>I am currently trying to get proteins out of inclusion bodies in E.coli
>after an overnight culture growth and am really battling. Anyone out
>there with advice to get prots out of inclusion bodies without
>destroying/denaturing the proteins and rendering them inactive?
>>Looking for help
>>Dr L.Davids, Cape Town, South Africa
If your protein is in an inclusion body it's already likely to be denatured as
it's not in a soluble form.
The only way to get it out is to solubilise appropriately (we use 6M Gd.HCl,
Tris buffer, pH 7, 10mM 2-ME) and then slowly remove the denaturing
extractants by dialysis.
Fingers crossed that the protein then refolds as you want it!
Cheers,
Roger
------------------------------------------------------------
Roger Murphy, Ph.D.
Biological Production Facility
Ludwig Institute for Cancer Research
Austin & Repatriation Medical Centre
Studley Road,
Heidelberg, Vic. 3084
Australia.
Tel 61-3-94965463
Fax 61-3-94965436
Email Roger.Murphy at Ludwig.edu.au
