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Prion diseases are fatal neurodegenerative disorders associated with the polymerization of the cellular form of prion protein (PrPC) into an amyloidogenic β-sheet infectious form (PrPSc). The sequence of host PrP is the major determinant of host prion disease susceptibility. In mice, the presence of allele a (Prnpa, encoding the polymorphism Leu-108/Thr-189) or b (Prnpb, Phe-108/Val-189) is associated with short or long incubation times, respectively, following infection with PrPSc. The molecular bases linking PrP sequence, infection susceptibility, and convertibility of PrPC into PrPSc remain unclear. Here we show that recombinant PrPa and PrPb aggregate and respond to seeding differently in vitro. Our kinetic studies reveal differences during the nucleation phase of the aggregation process, where PrPb exhibits a longer lag phase that cannot be completely eliminated by seeding the reaction with preformed fibrils. Additionally, PrPb is more prone to propagate features of the seeds, as demonstrated by conformational stability and electron microscopy studies of the formed fibrils. We propose a model of polymerization to explain how the polymorphisms at positions 108 and 189 produce the phenotypes seen in vivo. This model also provides insight into phenomena such as species barrier and prion strain generation, two phenomena also influenced by the primary structure of PrP. [ABSTRACT FROM AUTHOR]

Prions are lipidated proteins that interact with endogenous lipids and metal ions. They also assemble into multimers and propagate into the infectious scrapie form known as PrPSc. The high-resolution structure of the infectious PrPSc state remains unknown, and its analysis largely relies on detergent-based preparations devoid of endogenous ligands. Here we designed polymers that allow isolation of endogenous membrane:protein assemblies in native nanodiscs without exposure to conventional detergents that destabilize protein structures and induce fibrillization. A set of styrene-maleic acid (SMA) polymers including a methylamine derivative facilitated gentle release of the infectious complexes for resolution of multimers, and a thiol-containing version promoted crystallization. Polymer extraction from brain homogenates from Syrian hamsters infected with Hyper prions and WT mice infected with Rocky Mountain Laboratories prions yielded infectious prion nanoparticles including oligomers and microfilaments bound to lipid vesicles. Lipid analysis revealed the brain phospholipids that associate with prion protofilaments, as well as those that are specifically enriched in prion assemblies captured by the methylamine-modified copolymer. A comparison of the infectivity of PrPSc attached toSMAlipid particles in mice and hamsters indicated that these amphipathic polymers offer a valuable tool for high-yield production of intact, detergent-free prions that retain in vivo activity. This native prion isolation method provides an avenue for producing relevant prion:lipid targets and potentially other proteins that form multimeric assemblies and fibrils on membranes. [ABSTRACT FROM AUTHOR]

Lewy bodies and Lewy neurites, neuropathological hallmarks of several neurological diseases, are mainly made of filamentous assemblies of α-synuclein. However, other macromolecules including Tau, ubiquitin, glyceraldehyde-3-phosphate dehydrogenase, and glycosaminoglycans are routinely found associated with these amyloid deposits. Glyceraldehyde-3-phosphate dehydrogenase is a glycolytic enzyme that can form fibrillar aggregates in the presence of acidic membranes, but its role in Parkinson disease is still unknown. In this work, the ability of heparin to trigger the amyloid aggregation of this protein at physiological conditions of pH and temperature is demonstrated by infrared and fluorescence spectroscopy, dynamic light scattering, small angle x-ray scattering, circular dichroism, and fluorescence microscopy. Aggregation proceeds through the formation of short rod-like oligomers, which elongates in one dimension. Heparan sulfate was also capable of inducing glyceraldehyde-3-phosphate dehydrogenase aggregation, but chondroitin sulfates A, B, and C together with dextran sulfate had a negligible effect. Aided with molecular docking simulations, a putative binding site on the protein is proposed providing a rational explanation for the structural specificity of heparin and heparan sulfate. Finally, it is demonstrated that in vitro the early oligomers present in the glyceraldehyde-3- phosphate dehydrogenase fibrillation pathway promote α-synuclein aggregation. Taking into account the toxicity of α-synuclein prefibrillar species, the heparin-induced glyceraldehyde- 3-phosphate dehydrogenase early oligomers might come in useful as a novel therapeutic strategy in Parkinson disease and other synucleinopathies. [ABSTRACT FROM AUTHOR]