Prion Protein Aggregation and FibrillogenesisIn Vitro
This chapter focuses on the structural conversion of natural and recombinant prion proteins in vitro. They key event in prion diseases is the conversion of the cellular prion protein (PrPC) into its disease causing isoform PrPSc. This conversion is repres
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Prion Protein Aggregation and Fibrillogenesis In Vitro Jan Stöhr
Abstract This chapter focuses on the structural conversion of natural and recombinant prion proteins in vitro. They key event in prion diseases is the conversion of the cellular prion protein (PrPC ) into its disease causing isoform PrPSc . This conversion is represented by a conformational change from an α-helical dominated isoform into the mostly β-sheeted PrPSc . Represented is an overview of in vitro conversion systems that result in β-structured recombinant prion proteins including the current achievements in the generation of synthetic mammalian prions as proof of the protein-only hypothesis. In addition to the conversion of recombinant PrP the chapter features a summary of the protein misfolding cyclic amplification (PMCA) technique which has gained enormous popularity in prion research. Given is a general overview about the technique itself and the broad spectrum of utilization as detection method for prions. The spontaneous generation of prions by the protein misfolding amplification (PMCA) are also discussed. Keywords Prion · PrPC · PrPSc · Protein misfolding amplification (PMCA)
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Introduction
Prions are the causative agents of a wide range of neurodegenerative diseases in humans and animals. The key molecular event in the formation of prions is the conformational transition of the cellular isoform of the prion protein (PrPC ) into the disease causing isoform PrPSc (Prusiner 2007). PrPC is a mostly α-helical structured protein that is expressed in neuronal tissue in all mammalian species (Watts and Westaway 2007). The protein itself carries several posttranslational modifications, i.e. glycosylations and is anchored to the cell membrane of neurons through a glycophosphatidylinositol (GPI) membrane anchor (Stahl et al. 1987). The structural conversion from PrPC to PrPSc can be initiated spontaneously, through mutations within the prion protein sequence or by an exogenous infection with PrPSc molecules. Several lines of evidence showed that the misfolded prion protein itself is sufficient J. Stöhr () Institute for Neurodegenerative Diseases, University of California, San Francisco, CA 94143-0518, USA e-mail: [email protected] J. R. Harris (ed.), Protein Aggregation and Fibrillogenesis in Cerebral and Systemic Amyloid Disease, Subcellular Biochemistry 65, DOI 10.1007/978-94-007-5416-4_5, © Springer Science+Business Media Dordrecht 2012
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to create the infectious entity which represents the central dogma in the so-called protein-only hypothesis (Colby and Prusiner 2011; Legname et al. 2004; Deleault et al. 2007; Makarava et al. 2010). The protein-only hypothesis states that the structural conversion on its own is sufficient to create an infectious protein conformer that can self replicate if the substrate, i.e. PrPC , is present in the organism. The infectious cycle is then created by a reaction where PrPSc represents the structural template for recruiting more prion proteins from the cellular to the PrPSc isoform. A
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