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Langeveld, J.P.M.; Jacobs, J.G.; Erkens, J. H. F.; Bossers, Alex; Zijderveld, F.G. van; Keulen, L.J.M. van

doi:10.1186/1746-6148-2-19

Cited by 113 publications

(60 citation statements)

References 70 publications

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“…12B2 is classically used to differentiate type 1 and type 2 prions, and the 8G8 epitope is just downstream (Fig. 7A) (42,58). The lower band was negative for these antibodies but still positive for 8F9 that recognizes the C terminus of PrP C (Fig.…”

Section: Biochemical Characteristics Of Prpmentioning

confidence: 99%

Generating Bona Fide Mammalian Prions with Internal Deletions

Muñoz-Montesino

Sizun

Moudjou

et al. 2016

J Virol

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Mammalian prions are PrP proteins with altered structures causing transmissible fatal neurodegenerative diseases. They are self-perpetuating through formation of beta-sheet-rich assemblies that seed conformational change of cellular PrP. Pathological PrP usually forms an insoluble protease-resistant core exhibiting beta-sheet structures but no more alpha-helical content, loosing the three alpha-helices contained in the correctly folded PrP. The lack of a high-resolution prion structure makes it difficult to understand the dynamics of conversion and to identify elements of the protein involved in this process. To determine whether completeness of residues within the protease-resistant domain is required for prions, we performed serial deletions in the helix H2 C terminus of ovine PrP, since this region has previously shown some tolerance to sequence changes without preventing prion replication. Deletions of either four or five residues essentially preserved the overall PrP structure and mutant PrP expressed in RK13 cells were efficiently converted into bona fide prions upon challenge by three different prion strains. Remarkably, deletions in PrP facilitated the replication of two strains that otherwise do not replicate in this cellular context. Prions with internal deletion were self-propagating and de novo infectious for naive homologous and wild-type PrP-expressing cells. Moreover, they caused transmissible spongiform encephalopathies in mice, with similar biochemical signatures and neuropathologies other than the original strains. Prion convertibility and transfer of strain-specific information are thus preserved despite shortening of an alpha-helix in PrP and removal of residues within prions. These findings provide new insights into sequence/structure/infectivity relationship for prions. IMPORTANCEPrions are misfolded PrP proteins that convert the normal protein into a replicate of their own abnormal form. They are responsible for invariably fatal neurodegenerative disorders. Other aggregation-prone proteins appear to have a prion-like mode of expansion in brains, such as in Alzheimer's or Parkinson's diseases. To date, the resolution of prion structure remains elusive. Thus, to genetically define the landscape of regions critical for prion conversion, we tested the effect of short deletions. We found that, surprisingly, removal of a portion of PrP, the C terminus of alpha-helix H2, did not hamper prion formation but generated infectious agents with an internal deletion that showed characteristics essentially similar to those of original infecting strains. Thus, we demonstrate that completeness of the residues inside prions is not necessary for maintaining infectivity and the main strain-specific information, while reporting one of the few if not the only bona fide prions with an internal deletion.

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Section: Biochemical Characteristics Of Prpmentioning

confidence: 99%

Generating Bona Fide Mammalian Prions with Internal Deletions

Muñoz-Montesino

Sizun

Moudjou

et al. 2016

J Virol

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“…The following monoclonal mouse antibodies, immunoreactive with human PrP, were used: 3F4 at 1:30,000, which recognizes residues 106 to 110 (35); 12B2, at 1:8,000, which binds residues 89 to 93 (36); and SAF60 at 1:2,000, which reacts with residues 157 to 161 (37). In addition, the PrP Sc type 2-specific polyclonal antibody T2 (1:5,000), which binds residues 97 to 103 (7), and the rabbit antiserum 2301 (1: 3,000) to human PrP residues 220 to 231 were used.…”

Section: Patients and Tissuesmentioning

confidence: 99%

Analysis of Conformational Stability of Abnormal Prion Protein Aggregates across the Spectrum of Creutzfeldt-Jakob Disease Prions

Cescatti

Saverioni

Capellari

et al. 2016

J Virol

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The wide phenotypic variability of prion diseases is thought to depend on the interaction of a host genotype with prion strains that have self-perpetuating biological properties enciphered in distinct conformations of the misfolded prion protein PrP Sc . This concept is largely based on indirect approaches studying the effect of proteases or denaturing agents on the physicochemical properties of PrP Sc aggregates. Furthermore, most data come from studies on rodent-adapted prion strains, making current understanding of the molecular basis of strains and phenotypic variability in naturally occurring diseases, especially in humans, more limited. To fill this gap, we studied the effects of guanidine hydrochloride (GdnHCl) and heating on PrP Sc aggregates extracted from 60 sporadic Creutzfeldt-Jakob disease (CJD) and 6 variant CJD brains. While denaturation curves obtained after exposure of PrP Sc to increasing GdnHCl concentrations showed similar profiles among the 7 CJD types analyzed, PrP Sc exposure to increasing temperature revealed significantly different and type-specific responses. In particular, MM1 and VV2, the most prevalent and fast-replicating CJD types, showed stable and highly resistant PrP Sc aggregates, whereas VV1, a rare and slowly propagating type, revealed unstable aggregates that easily dissolved at low temperature. Taken together, our results indicate that the molecular interactions mediating the aggregation state of PrP Sc , possibly enciphering strain diversity, are differently targeted by GdnHCl, temperature, and proteases. Furthermore, the detected positive correlation between the thermostability of PrP Sc aggregates and disease transmission efficiency makes inconsistent the proposed hypothesis that a decrease in conformational stability of prions results in an increase in their replication efficiency. IMPORTANCEPrion strains are defined as infectious isolates propagating distinctive phenotypic traits after transmission to syngeneic hosts. Although the molecular basis of prion strains is not fully understood, it is largely accepted that variations in prion protein conformation drive the molecular changes leading to the different phenotypes. In this study, we exposed abnormal prion protein aggregates encompassing the spectrum of human prion strains to both guanidine hydrochloride and thermal unfolding. Remarkably, while exposure to increasing temperature revealed significant strain-specific differences in the denaturation profile of the protein, treatment with guanidine hydrochloride did not. The findings suggest that thermal and chemical denaturation perturb the structure of prion protein aggregates differently. Moreover, since the most thermostable prion protein types were those associated with the most prevalent phenotypes and most rapidly and efficiently transmitting strains, the results suggest a direct correlation between strain replication efficiency and the thermostability of prion protein aggregates. P rion diseases are invariably fatal neurodegenerative disorders of humans ...

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“…In addition, two rabbit antiserums against either PrP N terminus (residues [23][24][25][26][27][28][29][30][31][32][33][34][35][36][37][38][39][40] or PrP C terminus (the 2301 antiserum, residues 220 -231) were used.…”

Section: Antibodiesmentioning

confidence: 99%

Characterization of Truncated Forms of Abnormal Prion Protein in Creutzfeldt-Jakob Disease

Notari

Strammiello

Capellari

et al. 2008

Journal of Biological Chemistry

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In prion disease, the abnormal conformer of the cellular prion protein, PrP Sc , deposits in fibrillar protein aggregates in brain and other organs. Limited exposure of PrP Sc to proteolytic digestion in vitro generates a core fragment of 19 -21 kDa, named PrP27-30, which is also found in vivo. Recent evidence indicates that abnormal truncated fragments other than PrP27-30 may form in prion disease either in vivo or in vitro. We characterized a novel protease-resistant PrP fragment migrating 2-3 kDa faster than PrP27-30 in Creutzfeldt-Jakob disease (CJD) brains. The fragment has a size of about 18.5 kDa when associated with PrP27-30 type 1 (21 kDa) and of 17 kDa when associated with type 2 (19 kDa). Molecular mass and epitope mapping showed that the two fragments share the primary N-terminal sequence with PrP27-30 types 1 and 2, respectively, but lack a few amino acids at the very end of C terminus together with the glycosylphosphatidylinositol anchor. The amounts of the 18.5-or 17-kDa fragments and the previously described 13-kDa PrP Sc C-terminal fragment relatively to the PrP27-30 signal significantly differed among CJD subtypes.

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Cited by 113 publications

References 70 publications

Generating Bona Fide Mammalian Prions with Internal Deletions

Generating Bona Fide Mammalian Prions with Internal Deletions

Analysis of Conformational Stability of Abnormal Prion Protein Aggregates across the Spectrum of Creutzfeldt-Jakob Disease Prions

Characterization of Truncated Forms of Abnormal Prion Protein in Creutzfeldt-Jakob Disease

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