| Description |
Aβ(1-40) is the major fibril-forming peptide from Alzheimer’s disease. Monomeric Aβ is unstructured but 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, these fibrils are the products of a complex formation mechanism that 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 because binding of the antibody B10AP prevents the conversion of these metastable intermediates into mature fibrils. A set of eight peptides with varying isotope labeling schemes was obtained from chemical synthesis. The labels cover 30 residues that are distributed over the entire peptide sequence. 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 the combined data from chemical shifts and TALOS, 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 encompasses residues 23-26 and forms an intermediate segment in between the adjacent β-strands. The other two regions occur at the peptide N-terminus and within a small C-terminal segment. 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. We find that 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.
Further, we have carried solid-state NMR experiments on the microtubuli binding domain of the human tau protein, which is also involved in Alzheimers disease. Our experiments show that K19 binds to negatively charged membranes with high affinity thereby altering the lipid headgroup orientation and dynamics. In contrast, the lipid chain region is not influenced. K19 forms a β-sheet secondary structure on the membrane surface, however, no amyloid fibrils could be found.
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