Polymorphism and Ultrastructural Organization of Prion Protein Amyloid Fibrils: An Insight from High Resolution Atomic Force Microscopy

Anderson, Maighdlin; Bocharova, Olga V.; Makarava, Natallia; Breydo, Leonid; Salnikov, Vadim V.; Baskakov, Ilia V.

doi:10.1016/j.jmb.2006.02.007

Cited by 114 publications

(134 citation statements)

References 68 publications

(82 reference statements)

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“…1). These fibrils were morphologically similar to those produced from WT PrP (25). Taken together, these data confirmed that the incorporation of acrylodan-labeled Cys-PrPs into fibrils did not have noticeable effects on fibrillar secondary structure, the size of the PK-resistant core, or the ability of filaments to assemble into mature high-order fibrils.…”

Section: Resultssupporting

confidence: 74%

Site-specific Conformational Studies of Prion Protein (PrP) Amyloid Fibrils Revealed Two Cooperative Folding Domains within Amyloid Structure

Sun

Breydo

Makarava

et al. 2007

Journal of Biological Chemistry

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Despite the ability of most proteins to form amyloid, very little is know about amyloid fibril structures and the factors that govern their stability. Using amyloid fibrils produced from fulllength prion protein (PrP), we describe a reliable approach for determining both site-specific and global conformational stability of the fibrillar form. To measure site-specific stability, we produced six variants of PrP by replacing the residues at positions 88, 98, 127, 144, 196, and 230 with cysteine, labeled the new cysteines with the fluorescent dye acrylodan, and investigated their conformational status within the amyloid form in guanidine hydrochloride-induced denaturation experiments. We found that the fibrils labeled at positions 127, 144, 196, and 230 displayed cooperative unfolding and showed a very high C1 ⁄ 2 value similar to that observed for the global unfolding of the amyloid structure. The unfolding at residue 98 was also cooperative; however, it showed a C1 ⁄ 2 value substantially lower than that of global unfolding, whereas the unfolding of fibrils labeled at residue 88 was non-cooperative. These data illustrate that there are at least two independent cooperative folding domains within the amyloid structure of the full-length PrP. In addition, kinetic experiments revealed only a partial overlap between the region that constituted the fibrillar cross-␤ core and the regions that were involved in nucleation. This result illustrates that separate PrP regions accounted for the nucleation and for the formation of the conformationally most stable fibrillar core.

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Section: Resultssupporting

confidence: 74%

Site-specific Conformational Studies of Prion Protein (PrP) Amyloid Fibrils Revealed Two Cooperative Folding Domains within Amyloid Structure

Sun

Breydo

Makarava

et al. 2007

Journal of Biological Chemistry

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“…Collected images were processed with WCIF ImageJ software (National Institutes of Health) as described previously (12). Atomic force microscopy (AFM), negative staining electron microscopy (EM), and Fourier transform infrared spectroscopy (FTIR) were performed as described in our former publications (13,14).…”

Section: Methodsmentioning

confidence: 99%

“…S1). It is likely that the polymorphism within fibrils of R-or S-types is attributed to the variable number of constitutive protofilaments and variable modes of lateral association of protofilaments within individual fibrils (13,15). Regardless of the polymorphism observed within each individual class, S-and R-fibrils displayed remarkable differences with respect to their substructures, as described below.…”

Section: Prp Forms Structurally Different Fibrils Under Different Shamentioning

confidence: 97%

The Same Primary Structure of the Prion Protein Yields Two Distinct Self-propagating States

Makarava

Baskakov

2008

Journal of Biological Chemistry

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The question of whether distinct self-propagating structures could be formed within the same amino acid sequence in the absence of external cofactors or templates has important implications for a number of issues, including the origin of prion strains and the engineering of smart, self-assembling peptidebased biomaterials. In the current study, we showed that chemically identical prion protein can give rise to conformationally distinct, self-propagating amyloid structures in the absence of cellular cofactors, post-translational modification, or PrP Scspecified templates. Even more surprising, two self-replicating states were produced under identical solvent conditions, but under different shaking modes. Individual prion conformations were inherited by daughter fibrils in seeding experiments conducted under alternative shaking modes, illustrating the high fidelity of fibrillation reactions. Our study showed that the ability to acquire conformationally different self-propagating structures is an intrinsic ability of protein fibrillation and strongly supports the hypothesis that conformational variation in selfpropagating protein states underlies prion strain diversity.The existence of multiple prion strains is considered to be one of the most puzzling features of prion biology. According to the protein-only hypothesis of prion propagation, the normal cellular isoform of the prion protein, PrP C , can be converted into a range of self-propagating disease-related structures, referred to as strains of PrP Sc (1). Although certain physical properties are common for all PrP Sc strains, each strain also possesses unique strain-specific conformational features. Within the protein-only hypothesis, it is assumed that variation in PrP Sc structure can give rise to a broad range of strain-specific disease phenotypes, including substantial variation in incubation time to disease, in pathology, and in behavioral symptoms.In the past several years considerable evidence has accumulated indicating that prion strains do, indeed, exhibit notable differences in the amount and type of ␤-sheet structure, conformational stability, proteolytic resistance, and surface-exposed epitopes (2-7). Although the results of these aforementioned studies were consistent with the protein-only hypothesis, the primary origin of strain-specific conformational variations remains unclear. What is the possible source of strain-specific structural variation in PrP Sc ? A "unified theory" of prion propagation postulates that the strain-specific features of the prion infectious agent may be encoded by a coprion, or small nucleic acid (8). Alternatively, the ability to adopt multiple self-propagating structures may be an intrinsic property of the fibrillation reaction itself and can be achieved in the absence of a coprion. In the past, two distinct amyloid states were produced from chemically identical A␤ peptides (9); however, this property has never been demonstrated for fibrillation of larger polypeptides or global proteins such as PrP.In the current study...

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“…The histogram in Figure 4C showing the fibril widths distribution illustrates that the fibril species have an average width of 30 § 11 nm, which is identical to literature values. 9 In contrast, ovine PrP Sc -fibrils Sc in living cells which was observed by light, atomic force, and scanning electron microscopy to form thin, 5 mm-long structures on the cell surface. 36,37 Regarding the average width of PrP Sc -fibrils, their appearance in our AFM images was obscured by co-purified brain matter in the form of extraneous non-fibrillar material.…”

Section: Secondary and Ultrastructural Propertiesmentioning

confidence: 99%

“…Studies have so far been limited to relatively low-resolution techniques. [6][7][8][9][10] These studies have suggested a superpleated b-sandwich model for recPrP-fibrils characterized by a parallel, in-register alignment of b-strands within a core domain comprising residues 160-220. 6 Alternatively, a lefthanded parallel b-helix with the b-sheet core between residues 89-175 has been suggested based on EM studies on 2D-crystals of PrP27-30, a fragment of PrP Sc resulting from proteinase Kdigestion of the »90 N-terminal residues.…”

Section: Introductionmentioning

confidence: 99%

Progress towards structural understanding of infectious sheep PrP-amyloid

Müller

Brener

Andréoletti

et al. 2014

Prion

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The still elusive structural difference of non-infectious and infectious amyloid of the mammalian prion protein (PrP) is a major pending milestone in understanding protein-mediated infectivity in neurodegenerative diseases. Preparations of PrP-amyloid proven to be infectious have never been investigated with a high-resolution technique. All available models to date have been based on low-resolution data. Here, we establish protocols for the preparation of infectious samples of full-length recombinant (rec) PrP-amyloid in NMR-sufficient amounts by spontaneous fibrillation and seeded fibril growth from brain extract. We link biological and structural data of infectious recPrP-amyloid, derived from bioassays, atomic force microscopy, and solid-state NMR spectroscopy. Our data indicate a semi-mobile N-terminus, some residues with secondary chemical shifts typical of a-helical secondary structure in the middle part between »115 to »155, and a distinct b-sheet core C-terminal of residue »155. These findings are not in agreement with all current models for PrP-amyloid. We also provide evidence that samples seeded from brain extract may not differ in the overall arrangement of secondary structure elements, but rather in the flexibility of protein segments outside the b-core region. Taken together, our protocols provide an essential basis for the high-resolution characterization of non-infectious and infectious PrP-amyloid in the near future.

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Polymorphism and Ultrastructural Organization of Prion Protein Amyloid Fibrils: An Insight from High Resolution Atomic Force Microscopy

Cited by 114 publications

References 68 publications

Site-specific Conformational Studies of Prion Protein (PrP) Amyloid Fibrils Revealed Two Cooperative Folding Domains within Amyloid Structure

Site-specific Conformational Studies of Prion Protein (PrP) Amyloid Fibrils Revealed Two Cooperative Folding Domains within Amyloid Structure

The Same Primary Structure of the Prion Protein Yields Two Distinct Self-propagating States

Progress towards structural understanding of infectious sheep PrP-amyloid

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