PrP^Sc-Specific Antibody Reveals C-Terminal Conformational Differences between Prion Strains

Abstract

Understanding the structure of PrPSc and its strain variation has been one of the major challenges in prion disease biology. To study the strain-dependent conformations of PrP Sc , we purified proteinase-resistant PrP Sc (PrP RES ) from mouse brains with three different murine-adapted scrapie strains (Chandler, 22L, and Me7) and systematically tested the accessibility of epitopes of a wide range of anti-PrP and anti-PrP Sc specific antibodies by indirect enzyme-linked immunosorbent assay (ELISA). We found that epitopes of most anti-PrP antibodies were hidden in the folded structure of PrP RES , even though these epitopes are revealed with guanidine denaturation. However, reactivities to a PrP Sc -specific conformational C-terminal antibody showed significant differences among the three different prion strains. Our results provide evidence for strain-dependent conformational variation near the C termini of molecules within PrP Sc multimers. IMPORTANCEIt has long been apparent that prion strains can have different conformations near the N terminus of the PrP Sc protease-resistant core. Here, we show that a C-terminal conformational PrP Sc -specific antibody reacts differently to three murine-adapted scrapie strains. These results suggest, in turn, that conformational differences in the C terminus of PrP Sc also contribute to the phenotypic distinction between prion strains. T ransmissible spongiform encephalopathies (TSEs), or prion diseases, are fatal neurodegenerative diseases resulting in the accumulation of the misfolded form of prion protein (PrP) in the brain (1). Prions are disease-causing infectious agents that lack agent-coding nucleic acids (1). The normal cellular glycoprotein PrP (PrP C ), which is typically bound to a carboxyl-terminal glycosylphosphatidylinositol (GPI) anchor, can undergo major conformational changes into pathogenic disease-causing forms of PrP (PrP Sc ). This conversion is induced by the binding and templating effects of preexisting PrP Sc (2). Relative to PrP C , PrP Sc tends to be rich in ␤-sheets, detergent insoluble, oligomeric or fibrillar, and partially resistant to proteinase K (PK) digestion. PK treatment of PrP Sc typically produces a PK-resistant carboxyl-terminal core referred to as PrP RES or PrP(27-30) (3, 4). Although there is increasing evidence that protease-sensitive forms of disease-associated PrP can also exist in the brains of humans and animals affected with prion diseases (5-8), the presence of PrP RES is a major diagnostic indicator of prion diseases. However, the detailed threedimensional structures of PrP Sc and its variants remain a mystery. One approach to probing prion structures and studying prion pathogenesis has been the development of PrP Sc -and/or PrP RESselective antibodies (9-21). Despite some successes, the development of PrP Sc -specific antibodies with diverse epitopes has been limited by the fact that PrP Sc has the same primary sequence as PrP C . This requires PrP Sc -specific epitopes to be conformational. However, such potentially...

Disulfide-crosslink scanning reveals prion–induced conformational changes and prion strain–specific structures of the pathological prion protein PrPSc

“…This finding is also consistent with the strain–specific resistance of mice expressing PrP with N170S ( 37 ), implying a strain–dependent impact of regional structures in H1∼H2. Besides, differences between ME7, 22L, and RML prion strains in the regional structures of PrP Sc in the Ctrm region are also implied by their distinct immunoreactivity ( 38 ). Our proposed model specifically exemplifies regional structures that could explain those observations.…”

Mammalian amyloidogenic proteins promote prion nucleation in yeast

“…While the structural organization of PrP in a prion form remains a matter of debates (e. g., [66,67,68], our data agree with models locating cross-β interactions within the region 90-170, suggesting the retention of the native secondary structure by the C-terminal region of PrP (e. g., [68,69,70], and predict that the proposed β-structure at positions 160-164 is dispensable for prion initiation. However, our data do not necessarily contradict a possibility of further expansion of the amyloid core to the C-proximal region as shown for some PrP-based amyloids [71,72]. Most importantly, our yeast assay provides a tool that could be employed to further decipher sequential and structural requirements for initiation of PrP polymerization and conformational conversion.…”

Cofactor and glycosylation preferences for in vitro prion conversion are predominantly determined by strain conformation

“…Based on our findings, we suggest that the mechanism of strain-dependent neurotropism depends cofactor and PrP C glycoform selection by the template PrP Sc conformer during self-replication. This model explains how neurotropism and other biological strain properties can be associated with specific PrP Sc conformations [58][59][60][61][62][63][64]. It is intriguing to consider the role that cofactor molecules and post-translational modifications may play in the specific regional distribution patterns seen in other neurodegenerative…”