Hot spots in prion protein for pathogenic conversion (original) (raw)
Related papers
Accounts of Chemical Research, 2006
No validated treatments exist for transmissible spongiform encephalopathies (TSEs or prion diseases) in humans or livestock. The search for TSE therapeutics is complicated by persistent uncertainties about the nature of mammalian prions and their pathogenic mechanisms. In pursuit of anti-TSE drugs, we and others have focused primarily on blocking conversion of normal prion protein, PrP(C), to the TSE-associated isoform, PrP(Sc). Recently developed high-throughput screens have hastened the identification of new inhibitors with strong in vivo anti-TSE activities such as porphyrins, phthalocyanines, and phosphorthioated oligonucleotides. New routes of administration have enhanced beneficial effects against established brain infections. Several different classes of TSE inhibitors share structural similarities, compete for the same site(s) on PrP(C), and induce the clustering and internalization of PrP(C) from the cell surface. These activities may represent a common mechanism of action for these anti-TSE compounds.
A conformational switch controlling the toxicity of the prion protein
2021
SummaryPrion infections cause conformational changes of PrPC and lead to progressive neurological impairment. Here we show that toxic, prion-mimetic ligands induce an intramolecular R208-H140 hydrogen bond (“H-latch”) altering the flexibility of the α2-α3 and β2-α2 loops of PrPC. Expression of a PrP2Cys mutant mimicking the H-latch was constitutively toxic, whereas a PrPR207A mutant unable to form the H-latch conferred resistance to prion infection. High-affinity ligands that prevented H-latch induction repressed prion-related neurodegeneration in organotypic cerebellar cultures. We then selected phage-displayed ligands binding wild-type PrPC, but not PrP2Cys. These binders depopulated H-latched conformers and conferred protection against prion toxicity. Finally, brain-specific expression of an antibody rationally designed to prevent H-latch formation, prolonged the life of prion-infected mice despite unhampered prion propagation, confirming that the H-latch is causally linked to pr...
Discovery of Novel Anti-prion Compounds Using In Silico and In Vitro Approaches
Prion diseases are associated with the conformational conversion of the physiological form of cellular prion protein (PrP C) to the pathogenic form, PrP Sc. Compounds that inhibit this process by blocking conversion to the PrP Sc could provide useful anti-prion therapies. However, no suitable drugs have been identified to date. To identify novel anti-prion compounds, we developed a combined structureand ligand-based virtual screening system in silico. Virtual screening of a 700,000-compound database, followed by cluster analysis, identified 37 compounds with strong interactions with essential hotspot PrP residues identified in a previous study of PrP C interaction with a known antiprion compound (GN8). These compounds were tested in vitro using a multimer detection system, cell-based assays, and surface plasmon resonance. Some compounds effectively reduced PrP Sc levels and one of these compounds also showed a high binding affinity for PrP C. These results provide a promising starting point for the development of anti-prion compounds. Prion diseases are a group of lethal neurodegenerative diseases of humans and animals, including human Creutzfeldt-Jakob disease; bovine spongiform encephalopathy; scrapie in sheep, hamsters, and mice; and chronic wasting diseases in deer 1,2. There are three causes of prion disease: hereditary, sporadic, and acquired by infection. All of these disease types are known to share the same pathogenic mechanism 2,3. The central event in prion disease pathogenesis is the conversion of the α-helix-rich cellular form of prion protein (PrP C) to a misfolded, β-sheet-rich, pathogenic, and infectious conformational isoform (PrP Sc), although the detailed structure of PrP Sc is still not fully characterised 1,4,5. This conversion initiates a chain replication reaction, where each newly converted PrP Sc molecule interacts with more PrP C molecules, fueling the formation of additional PrP Sc6,7. After this post-translational conversion, PrP Sc aggregates and becomes the detergent-insoluble, partially protease-resistant protein fraction that serves as the marker for prion diseases 8,9. Therefore, stabilization of the native PrP C conformation, without blocking the normal functions of PrP C , could reduce the rate of conversion to PrP Sc or even prevent prion disease. To date, screening has led to the identification of many anti-prion compounds 10. Several large molecules (pentosanpolysulfate 5 , suramin 11 , amphotericin B 12 , congo red 13 , and dendritic polyamines 14) and small molecules (bis-acridine 15 , polyphenol, phenothiazine, anti-histamine, statin, and some anti-malarial agents including quinacrine 16) have been reported to inhibit PrP Sc formation or to reduce the level of PrP C. The tyrosine kinase inhibitor, STI571 (Gleevec), cured scrapie-infected cells in a concentration-and
Antimicrobial agents and chemotherapy, 2009
Transmissible spongiform encephalopathies are associated with the conformational conversion of the prion protein from the cellular form (PrP(C)) to the scrapie form. This process could be disrupted by stabilizing the PrP(C) conformation, using a specific ligand identified as a chemical chaperone. To discover such compounds, we employed an in silico screen that was based on the nuclear magnetic resonance structure of PrP(C). In combination, we performed ex vivo screening using the Fukuoka-1 strain-infected neuronal mouse cell line at a compound concentration of 10 microM and surface plasmon resonance. Initially, we selected 590 compounds according to the calculated docked energy and finally discovered 24 efficient antiprion compounds, whose chemical structures are quite diverse. Surface plasmon resonance studies showed that the binding affinities of compounds for PrP(C) roughly correlated with the compounds' antiprion activities, indicating that the identification of chemical cha...
Journal of Neuroscience, 2011
Transmissible spongiform encephalopathies (TSEs) are fatal neurodegenerative diseases attributed to misfolding of the cellular prion protein, PrP C , into a β-sheet-rich, aggregated isoform, PrP Sc . We previously found that expression of mouse PrP with the two amino acid substitutions S170N and N174T, which result in high structural order of the β2-α2 loop in the NMR structure at pH 4.5 and 20 °C, caused transmissible de novo prion disease in transgenic mice. Here we report that expression of mouse PrP with the single-residue substitution D167S, which also results in a structurally well-ordered β2-α2 loop at 20 °C, elicits spontaneous PrP aggregation in vivo. Transgenic mice expressing PrP D167S developed a progressive encephalopathy characterized by abundant PrP plaque formation, spongiform change, and gliosis. These results add to the evidence that the β2-α2 loop has an important role in intermolecular interactions, including that it may be a key determinant of prion protein aggregation.
2011
Transmissible spongiform encephalopathies (TSEs) are fatal neurodegenerative diseases attributed to misfolding of the cellular prion protein, PrP C , into a β-sheet-rich, aggregated isoform, PrP Sc. We previously found that expression of mouse PrP with the two amino acid substitutions S170N and N174T, which result in high structural order of the β2-α2 loop in the NMR structure at pH 4.5 and 20 °C, caused transmissible de novo prion disease in transgenic mice. Here we report that expression of mouse PrP with the single-residue substitution D167S, which also results in a structurally well-ordered β2-α2 loop at 20 °C, elicits spontaneous PrP aggregation in vivo. Transgenic mice expressing PrP D167S developed a progressive encephalopathy characterized by abundant PrP plaque formation, spongiform change, and gliosis. These results add to the evidence that the β2-α2 loop has an important role in intermolecular interactions, including that it may be a key determinant of prion protein aggregation.
Structural attributes of mammalian prion infectivity: Insights from studies with synthetic prions
Journal of Biological Chemistry, 2018
Prion diseases are neurodegenerative disorders that affect many mammalian species. Mammalian prion proteins (PrPs) can misfold into many different aggregates. However, only a small subpopulation of these structures is infectious. One of the major unresolved questions in prion research is identifying which specific structural features of these misfolded protein aggregates are important for prion infectivity in vivo. Previously, two types of proteinase K-resistant, self-propagating aggregates were generated from the recombinant mouse prion protein in the presence of identical cofactors. Although these two aggregates appear biochemically very similar, they have dramatically different biological properties, with one of them being highly infectious and the other one lacking any infectivity. Here, we used several MS-based structural methods, including hydrogen-deuterium exchange and hydroxyl radical footprinting, to gain insight into the nature of structural differences between these two PrP aggregate types. Our experiments revealed a number of specific differences in the structure of infectious and noninfectious aggregates, both at the level of the polypeptide backbone and quaternary packing arrangement. In particular, we observed that a high degree of order and stability of -sheet structure within the entire region between residues ϳ89 and 227 is a primary attribute of infectious PrP aggregates examined in this study. By contrast, noninfectious PrP aggregates are characterized by markedly less ordered structure up to residue ϳ167. The structural constraints reported here should facilitate development of experimentally based high-resolution structural models of infectiosus mammalian prions.
Thermodynamic stabilization of the folded domain of prion protein inhibits prion infection in vivo
Cell reports, 2013
Prion diseases, or transmissible spongiform encephalopathies (TSEs), are associated with the conformational conversion of the cellular prion protein, PrP(C), into a protease-resistant form, PrP(Sc). Here, we show that mutation-induced thermodynamic stabilization of the folded, α-helical domain of PrP(C) has a dramatic inhibitory effect on the conformational conversion of prion protein in vitro, as well as on the propagation of TSE disease in vivo. Transgenic mice expressing a human prion protein variant with increased thermodynamic stability were found to be much more resistant to infection with the TSE agent than those expressing wild-type human prion protein, in both the primary passage and three subsequent subpassages. These findings not only provide a line of evidence in support of the protein-only model of TSEs but also yield insight into the molecular nature of the PrP(C)→PrP(Sc) conformational transition, and they suggest an approach to the treatment of prion diseases.