Purification and Refolding to Amyloid Fibrils of (His)6-tagged Recombinant Shadoo Protein Expressed as Inclusion Bodies in XXXX .

@entity1133 @entity8304 @entity1579 @entity179 @entity3411 @entity526
UNASSIGNED: The @entity526 expression system is a powerful tool for the production of recombinant eukaryotic proteins. We use it to produce Shadoo, a protein belonging to the prion family. A chromatographic method for the purification of (His)6-tagged recombinant Shadoo expressed as inclusion bodies is described. The inclusion bodies are solubilized in 8 M @entity1133 and bound to a @entity3411 -charged column to perform ion affinity chromatography. Bound proteins are eluted by a gradient of @entity8304 . Fractions containing Shadoo protein are subjected to size exclusion chromatography to obtain a highly purified protein. In the final step purified Shadoo is desalted to remove salts, @entity1133 and @entity8304 . Recombinant Shadoo protein is an important reagent for biophysical and biochemical studies of protein conformation disorders occurring in @entity1579 . Many reports demonstrated that prion @entity179 originate from the deposition of stable, ordered amyloid fibrils. Sample protocols describing how to fibrillate Shadoo into amyloid fibrils at acidic and neutral/basic pHs are presented. The methods on how to produce and fibrillate Shadoo can facilitate research in laboratories working on @entity1579 , since it allows for production of large amounts of protein in a rapid and low cost manner.
