The N-Terminal Sequence of Prion Protein Consists an Epitope Specific to the Abnormal Isoform of Prion Protein (PrPSc)

Masujin, Kentaro; Kaku-Ushiki, Yuko; Miwa, Ritsuko; Okada, Hiroyuki; Shimizu, Yoshihisa; Kasai, Kazuo; Matsuura, Yuichi; Yokoyama, Takashi

doi:10.1371/journal.pone.0058013

Cited by 11 publications

(11 citation statements)

References 48 publications

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“…e,f) could allow for selection of the desired mAbs. On the other hand, in the epitope mapping of 8D5 and 6A12, PrP Sc ‐specific mAbs recently generated by our group, Pepspot revealed that these mAbs recognize almost the same region, that is, PrP31–39 and PrP41–47, respectively . This finding supports the idea that the N‐terminus of PrP Sc harbors a PrP Sc ‐specific conformation.…”

Section: Evaluation Of Antisera Binding To the N‐terminal Region By Esupporting

confidence: 75%

Different antigenicities of the N‐terminal region of cellular and scrapie prion proteins

Ushiki-Kaku

Iwamaru

Masujin

et al. 2013

Microbiology and Immunology

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Limited information is available about conformational differences between the abnormal isoform of prion protein (PrP Sc ) and cellular prion protein (PrP C ) under native conditions. To clarify conformational differences between these two isoforms, PrP-deficient mice were immunized with brain homogenates of normal and scrapie-infected animals. All mice generated anti-PrP antibodies. Peptide array analysis of these serum samples revealed a distinctive epitope of PrP Sc consisting of (2). Nuclear magnetic resonance analysis of rPrP has shown that the N-terminal segment, PrP23-120, is highly flexible, unlike the core domain (3). Because PrP res retains transmissibility, the Nterminal region is not thought to contribute to prion pathogenesis (4). However, the N-terminal region of native PrP Sc has not been well studied and information on the conformation of this region is lacking. To investigate the conformational characteristics of this region, it is necessary to prepare native, intact PrP Sc . Most of the available anti-PrP mAbs, which have been generated by immunization with rPrP or synthetic peptides of PrP, react to both PrP C and PrP Sc (5, 6); there are a few exceptions (7,8). Anti-PrP res mAbs also react with both PrP C and PrP Sc , even when the immunogens have been purified from prion-infected animals (5, 9). Recently, several PrP Sc -specific mAbs have been generated by immunization with purified intact PrP Sc (10-13). These findings indicate that intact PrP Sc retains a specific conformation that may support its use as an immunogen for generation of PrP Scspecific mAbs.

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Section: Evaluation Of Antisera Binding To the N‐terminal Region By Esupporting

confidence: 75%

Different antigenicities of the N‐terminal region of cellular and scrapie prion proteins

Ushiki-Kaku

Iwamaru

Masujin

et al. 2013

Microbiology and Immunology

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“…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)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(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 .…”

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confidence: 99%

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

Saijo

Hughson

Raymond

et al. 2016

J Virol

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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...

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“…Moreover, a novel PrP Sc -specific epitope has been recently reported in the region aa31-aa47 by using mAbs raised against intact PrP Sc complexes (Masujin et al 2013). Surprisingly, in our panel, mAb EB8 failed to promote prion clearance as efficiently as the other antibodies.…”

Section: Discussioncontrasting

confidence: 49%

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Supplemental Information 2: Peptides Used for Epitope Mapping

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Prion diseases are a group of fatal neurodegenerative disorders that affect humans and animals. They are characterized by the accumulation in the central nervous system of a pathological form of the host-encoded prion protein (PrP C). The prion protein is a membrane glycoprotein that consists of two domains: a globular, structured C-terminus and an unstructured N-terminus. The N-terminal part of the protein is involved in different functions in both health and disease. In the present work we discuss the production and biochemical characterization of a panel of four monoclonal antibodies (mAbs) against the distal N-terminus of PrP C using a well-established methodology based on the immunization of Prnp 0/0 mice. Additionally, we show their ability to block prion (PrP Sc) replication at nanomolar concentrations in a cell culture model of prion infection. These mAbs represent a promising tool for prion diagnostics and for studying the physiological role of the N-terminal domain of PrP C .

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The N-Terminal Sequence of Prion Protein Consists an Epitope Specific to the Abnormal Isoform of Prion Protein (PrPSc)

Cited by 11 publications

References 48 publications

Different antigenicities of the N‐terminal region of cellular and scrapie prion proteins

Different antigenicities of the N‐terminal region of cellular and scrapie prion proteins

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

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The N-Terminal Sequence of Prion Protein Consists an Epitope Specific to the Abnormal Isoform of Prion Protein (PrPSc)

Cited by 11 publications

References 48 publications

Different antigenicities of the N‐terminal region of cellular and scrapie prion proteins

Different antigenicities of the N‐terminal region of cellular and scrapie prion proteins

PrPSc-Specific Antibody Reveals C-Terminal Conformational Differences between Prion Strains

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