Abstract
(English)
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The activities during the 3-year duration of this project were focused primarily on the cloning, expression, purification and NMR structure determination of prion proteins from selected mammalian and non-mammalian species. Structure determinations have been completed and published for the wild-type human prion protein, 3 variants of the human prion protein, and the bovine, sheep and chicken prion proteins. The prion proteins from the pig, the elk and the turtle, a variant human prion protein with two disulfide bonds, and the human doppel protein have been cloned and expressed, and the NMR structure determinations are in an advanced stage.
The NMR structures of the recombinant human prion protein hPrP(23-230), and two C-terminal fragments thereof, hPrP(90-230) and hPrP(121-230), include a globular domain extending from residues 125-228, for which a detailed structure was obtained, and an N-terminal flexibly disordered 'tail'. The globular domain contains three a-helices comprising the residues 144-154, 173-194 and 200-228, and a short anti-parallel b-sheet comprising the residues 128-131 and 161-164. For the bovine prion protein, NMR structures were determined for the intact recombinant polypeptide chain bPrP(23-230), and a C-terminal fragment, bPrP(121-230). These molecules include a globular domain extending from residues 125-227, which is virtually identical to the globular domain in human PrP. To further investigate the basis and possible functional consequences of the near-identity of the globular domains in hPrP and bPrP, as well as subtle structural differences between these proteins and mouse PrP or Syrian hamster PrP, the NMR structures of three single-amino acid variants of the C-terminal domain of the human prion protein were determined. It was found that distinct local conformational variations could be related with the single-amino acid exchanges between the human protein and either the mouse or the Syrian hamster protein. These comparative studies are currently being expanded based on the aforementioned new structure determinations.
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