The architecture of amyloid-like peptide fibrils revealed by X-ray scattering, diffraction and electron microscopy.

Annette E Langkilde,Bente Vestergaard,Dmitri I Svergun,Louise C Serpell,Kyle L Morris

doi:10.1107/s1399004715001674

Abstract

Structural analysis of protein fibrillation is inherently challenging. Given the crucial role of fibrils in amyloid diseases, method advancement is urgently needed. A hybrid modelling approach is presented enabling detailed analysis of a highly ordered and hierarchically organized fibril of the GNNQQNY peptide fragment of a yeast prion protein. Data from small-angle X-ray solution scattering, fibre diffraction and electron microscopy are combined with existing high-resolution X-ray crystallographic structures to investigate the fibrillation process and the hierarchical fibril structure of the peptide fragment. The elongation of these fibrils proceeds without the accumulation of any detectable amount of intermediate oligomeric species, as is otherwise reported for, for example, glucagon, insulin and α-synuclein. Ribbons constituted of linearly arranged protofilaments are formed. An additional hierarchical layer is generated via the pairing of ribbons during fibril maturation. Based on the complementary data, a quasi-atomic resolution model of the protofilament peptide arrangement is suggested. The peptide structure appears in a β-sheet arrangement reminiscent of the β-zipper structures evident from high-resolution crystal structures, with specific differences in the relative peptide orientation. The complexity of protein fibrillation and structure emphasizes the need to use multiple complementary methods.

Highlights

The severity of several amyloid diseases underlines the importance of studying the structural aspects of protein amyloid fibrillation (Cecchi & Stefani, 2013; Knowles et al, 2014)
The estimated molecular weight (MW) and radius of gyration (Rg) are in agreement with those of a monomeric peptide, and the theoretical scattering curve calculated from the monomeric peptide fits the experimental data (Supplementary Fig. S2)
Dmax is obtained during the indirect Fourier transformation to the pair-distance distributions (Supplementary Fig. S3c) but, like Rg, can only be estimated with some uncertainty when the length of the fibrils surpasses the resolution of the small-angle X-ray scattering (SAXS) data

Summary

Introduction

The severity of several amyloid diseases underlines the importance of studying the structural aspects of protein amyloid fibrillation (Cecchi & Stefani, 2013; Knowles et al, 2014). In spite of more than a century of dedicated research efforts, how amyloid-like fibrils are formed remains elusive. This is mainly because fibrillation constitutes an inherent structural analytical challenge, since fibril formation proceeds via several equilibria between native and unfolded or refolded structures, oligomers, protofilaments and mature fibrils. Many fragments of amyloidogenic proteins as well as synthetic peptides can form amyloid-like fibrils in vitro. Compared with fibrils formed from full-length amyloidogenic proteins, such peptide fibrils exhibit relatively lower complexity and provide ideal model systems for structural analysis (see, for example, Balbirnie et al, 2001; Reches et al, 2002)

Methods

Results

Conclusion