Nucleation-dependent conformational conversion of the Y145Stop variant of human prion protein: Structural clues for prion propagation

Abstract

One of the most intriguing disease-related mutations in human prion protein (PrP) is the Tyr to Stop codon substitution at position 145. This mutation results in a Gerstmann-Straussler-Scheinkerlike disease with extensive PrP amyloid deposits in the brain. Here, we provide evidence for a spontaneous conversion of the recombinant polypeptide corresponding to the Y145Stop variant (huPrP23-144) from a monomeric unordered state to a fibrillar form. This conversion is characterized by a protein concentration-dependent lag phase and has characteristics of a nucleation-dependent polymerization. Atomic force microscopy shows that huPrP23-144 fibrils are characterized by an apparent periodicity along the long axis, with an average period of 20 nm. Fourier-transform infrared spectra indicate that the conversion is associated with formation of ␤-sheet structure. However, the infrared bands for huPrP23-144 are quite different from those for a synthetic peptide PrP106 -126, suggesting conformational non-equivalence of ␤-structures in the disease-associated Y145Stop variant and a frequently used short model peptide. To identify the region that is critical for the self-seeded assembly of huPrP23-144 amyloid, experiments were performed by using the recombinant polypeptides corresponding to prion protein fragments 23-114, 23-124, 23-134, 23-137, 23-139, and 23-141. Importantly, none of the fragments ending before residue 139 showed a propensity for conformational conversion to amyloid fibrils, indicating that residues within the 138 -141 region are essential for this conversion.

The Effect of Disease-associated Mutations on the Folding Pathway of Human Prion Protein

“…In these cases, the pathogenic process could potentially be facilitated by mutation-dependent changes in the amyloidogenic propensity of the unstructured PrP C region. The role of this region in prion protein conversion is in line with a number of observations (55,56), including the recent finding that a polypeptide corresponding to residues 23-144 of PrP undergoes a self-propagating conversion to the amyloid state (57).…”

Dynamics of a truncated prion protein, PrP(113–231), from 15N NMR relaxation: Order parameters calculated and slow conformational fluctuations localized to a distinct region

“…This fragment lacks residues from helix B and C, but still causes a familial prion disease. 54 Slow conformational fluctuations are localized to a distinct region within PrP C Slow motions that cause exchange broadening of NMR signals are typically because of large segmental motions rather than single hydrogen-bond rearrangements. Residues exhibiting exchange broadening sample alternative conformations in exchange with the native conformation on a milli-micro second timescale.…”

Molecular conformation and dynamics of the Y145Stop variant of human prion protein in amyloid fibrils

“…Recently, we and others have shown that certain aspects of this process can be conveniently studied by using the recombinant Y145Stop mutant of the human prion protein, huPrP23-144 (14,15). This C-truncated protein, associated with prion protein hereditary amyloidosis known as PrP cerebral amyloid angiopathy (16), undergoes very efficient autocatalytic conformational conversion under physiologically relevant buffer conditions (14,15); a similar reaction for recombinant full-length prion protein or PrP90-231 has been reported only in the presence of relatively high concentrations of chemical denaturants (17,18). Most importantly, using this experimentally tractable model, we were able to reproduce in vitro some of the most fundamental aspects of prion propagation, including the phenomena of ''species barrier'' and prion strains (19,20).…”