Solid-state NMR structure of a pathogenic fibril of full-length human [alpha]-synuclein

• Published in: Nature structural & molecular biology Vol. 23; no. 5; p. 409
• Main Authors: Tuttle, Marcus D, Comellas, Gemma, Nieuwkoop, Andrew J, Covell, Dustin J, Berthold, Deborah A, Kloepper, Kathryn D, Courtney, Joseph M, Kim, Jae K, Barclay, Alexander M, Kendall, Amy, Wan, William, Stubbs, Gerald, Schwieters, Charles D, Lee, Virginia M Y, George, Julia M, Rienstra, Chad M
• Format: Journal Article
• Language: English
• Published: New York Nature Publishing Group 01.05.2016
• Subjects: Data collection; Molecular biology; Neurons; NMR; Nuclear magnetic resonance; Parkinson's disease; Pathogens; Peptides; Residues; Sample preparation; Topology
• ISSN: 1545-9993; 1545-9985
• DOI: 10.1038/nsmb.3194

PD is pathologically characterized by Lewy bodies and Lewy neurites1, intracytoplasmic aggregates containing -synuclein (-syn) fibrils1. Exogenous -syn fibrils seed Lewy body and Lewy neuritelike inclusions in cell culture models2,3, and neuron-to-neuron -syn transmission propagates PD-like pathology4. Inoculation of pre-formed -syn fibrils into wild-type (WT) nontransgenic mice seeds aggregation of endogenous mouse -syn and reproduces key features of the neurodegenerative cascade5. Additionally, recent studies in rat models have established that -syn fibril strains cause distinct synucleinopathies with differing toxicity profiles6. Although secondary structures have been examined for several -syn fibril forms by solid-state NMR (SSNMR) spectroscopy710, to date there is no reported high-resolution 3D structure. The -syn monomer (14.5 kDa) is substantially larger than other amyloid peptides or proteins whose structures have been solved, such as HET-s (8.7 kDa)11, a 2-microglobulin fragment (2.5 kDa)12, amyloid- (A)(140) (4.3 kDa)1315 and A(142) (4.5 kDa)16. In addition to its large size, the highly repetitive secondary structure and residue-type degeneracy in -syn present major challenges for determination of high-resolution structures. To address these challenges, we performed a comprehensive structural study of an -syn fibril form previously reported by our group8,17,18. Using extensive sample preparation (six isotopically labeled samples), data collection (68 multidimensional spectra) and computational analysis (interpretation of >7,500 cross-peaks), we determined a single unique conformation in which the core residues are arranged in parallel, in-register -sheets with a Greek-key topology. We validated the structure with measurements of fibril width, intermolecular stacking and -sheet spacing by EM and X-ray fiber diffraction. These structural insights establish the basis for an improved understanding of -syn fibril nucleation, propagation and interactions with small molecules of potential utility for the diagnosis and treatment of PD.