Abstract
Prion is an infectious protein (PrP(Sc)) that is derived from a cellular glycoprotein (PrP(C)) through a conformational transition and associated with a group of prion diseases in animals and humans. Characterization of proteinase K (PK)-resistant PrP(Sc) by western blotting has been critical to diagnosis and understanding of prion diseases including Creutzfeldt-Jakob disease (CJD) and Gerstmann-Sträussler-Scheinker (GSS) disease in humans. However, formation as well as biochemical and biological properties of the glycoform-selective PrP(Sc) in variably protease-sensitive prionopathy (VPSPr) remain poorly understood. Here we reveal that formation of the ladder-like PrP(Sc) in VPSPr is a PK-dependent two-step process, which is enhanced by basic pH. Two sets of PrP(Sc) fragments can be identified with antibodies directed against an intermediate or a C-terminal domain of the protein. Moreover, antibodies directed against specific PrP glycoforms reveal faster electrophoretic migrations of PrP fragments mono-glycosylated at residue 181 and 197 in VPSPr than those in sporadic CJD (sCJD). Finally, RT-QuIC assay indicates that PrP(Sc)-seeding activity is lower and its lag time is longer in VPSPr than in sCJD. Our results suggest that the glycoform-selective PrP(Sc) in VPSPr is associated with altered glycosylation, resulting in different PK-truncation and aggregation seeding activity compared to PrP(Sc) in sCJD.