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Title: Biochemical analysis of PrPSc associated with the transmission of ovine prions
Author: Hopkins, L. J.
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2009
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The hypothesis tested in this thesis was that passage of ovine scrapie isolates in isogenic hosts is associated with the formation of PrPSc with distinct molecular properties, thereby providing a molecular basis for the discrimination of ovine prion strains. The molecular profile of PrPSc associated with the mouse-adapted sheep scrapie strains RML and ME7 was investigated. Western blot analysis, together with denaturant-dependent conformational stability analysis, revealed that PrPSc associated with RML and ME7 prion strains showed distinct biochemical and biophysical properties. PrPSc associated with RML and ME7 exhibited similar resistance to Proteinase K digestion but differed significantly in the glycoform profile of the resultant PrP27-30. In addition, ME7 PrPSc showed greater conformational stability than did RML PrPSc following denaturation with guanidine hydrochloride and proteolytic cleavage with Proteinase K. These novel data demonstrated that mouse-passaged ovine scrapie strains RML and ME7 could be discriminated at the molecular level by distinct conformers of PrPSc. Subsequently, PrPSc associated with primary passage, in conventional and ovine PrP transgenic mouse lines, of different isolates of natural ovine scrapie was investigated for the presence of distinct conformers of prion protein. Transmission of ARQ homozygous scrapie isolates resulted in the accumulation of significantly greater levels of PrPSc in Prnpa, Prnpb and ovine ARQ PrP transgenic mice than did VRQ homozygous inocula. In contrast, similar levels of PrPSc were generated in ovine VRQ transgenic mice. Significantly, the degree of conformational stability of the accumulated PrPSc correlated with its glycoform profile. transmission of ARQ homozygous scrapie isolates induced the accumulation of PrPSc characterised by a predominance of di-glycosylated PrPSc that was significantly more stable than that associated with the passage of VRQ homozygous inocula, which had similar levels of di- and mono-glycosylated PrPSc. These novel observations provide a biochemical basis for ovine prion strain typing.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available