Detection and characterization of proteinase K-sensitive disease-related prion protein with thermolysin

Sabrina Cronier (1) , Nathalie Gros , M. Howard Tattum , Graham S Jackson , Anthony R. Clarke , John Collinge , Jonathan D.F. Wadsworth

1 Department of Neurodegenerative Diseases

Disease-related prion protein, PrPSc, is classically distinguished from its normal cellular precursor, PrPC, by its detergent insolubility and partial resistance to proteolysis. Although molecular diagnosis of prion disease has historically relied upon detection of protease resistant fragments of PrPSc using proteinase K (PK), it is now apparent that a substantial fraction of disease-related PrP is destroyed by this protease. Recently, thermolysin has been identified as a complementary tool to PK, permitting isolation of PrPSc in its full-length form. Here we show that thermolysin can degrade PrPC while preserving both PK-sensitive and PK-resistant isoforms of disease-related PrP in both rodent and human prion strains. For mouse RML prions, the majority of PK-sensitive disease-related PrP isoforms do not appear to contribute significantly to infectivity. In variant Creutzfeldt-Jakob disease (vCJD), the human counterpart of bovine spongiform encephalopathy (BSE), up to 90 % of total PrP present in brain resists degradation with thermolysin whereas only ~15 % of this material resists digestion by PK. Detection of PK-sensitive isoforms of disease-related PrP using thermolysin should be useful for improving diagnostic sensitivity in human prion diseases.

Mots clés: Life Sciences

Sabrina Cronier, Nathalie Gros, M. Howard Tattum, Graham S Jackson, Anthony R. Clarke, et al.. Detection and characterization of proteinase K-sensitive disease-related prion protein with thermolysin. Biochemical Journal, 2008, 416 (2), pp.297-305. ⟨10.1042/BJ20081235⟩. ⟨hal-00479051⟩