A solid-State NMR Study of Abeta Protofibrils

Abstract

Aβ(1-40) is the major fibril-forming peptide from Alzheimer's disease, which adopts a highly ordered β-sheet conformation upon aggregation into amyloid fibrils. Several techniques have provided a wealth of structural information on mature Aβ(1-40) fibrils. Yet, the complex formation process of mature fibrils is, from a structural point of view, not well understood. Here we use ssNMR spectroscopy to elucidate the structure of Aβ protofibrils. This analysis is possible since the binding of the antibody B10AP prevents the conversion of these metastable intermediates into mature fibrils. With a set of eight peptides with varying isotope labeling schemes, 30 residues distributed over the entire peptide sequence were covered. 13C CPMAS spectra and 2D correlation experiments were recorded for unambiguous assignment of all carbons. From the conformation dependent chemical shifts we could identify peptide segments of stable secondary structure and evaluated the backbone structure using TALOS. Based on this data, Aβ protofibrils encompass residues 16-22 and 30-36 in β-sheet conformation. Further, three structural regions of the protofibrils present random coil-like chemical shifts, one (residues 23-26) as intermediate segment between the β-strands and two others at the peptide N- and C- terminus. Obviously the structural elements of mature Aβ(1-40) fibrils are already present in protofibrils, but the β-sheet regions apparently elongate during the fibrils conversion. Further information about the dynamics of these regions is provided by measurement of the strength of dipolar couplings, which are converted into an order parameter. Protofibrils show high order parameters (>0.8) within the β-strand regions, while the measured S values are below 0.8 at the termini. We never observed S values below 0.4 that would have indicated very high mobility. Thus, significant structural order exists also within those sequence segments that have chemical shift values corresponding to a random coil.