The role of fibril structure and surface hydrophobicity in secondary nucleation of amyloid fibrils

Abstract

Crystals, nanoparticles, and fibrils catalyze the generation of new aggregates on their surface from the same type of monomeric building blocks as the parent assemblies. This secondary nucleation process can be many orders of magnitude faster than primary nucleation. In the case of amyloid fibrils associated with Alzheimer’s disease, this process leads to the multiplication and propagation of aggregates, whereby short-lived oligomeric intermediates cause neurotoxicity. Understanding the catalytic activity is a fundamental goal in elucidating the molecular mechanisms of Alzheimer’s and associated diseases. Here we explore the role of fibril structure and hydrophobicity by asking whether the V18, A21, V40, and A42 side chains which are exposed on the Aβ42 fibril surface as continuous hydrophobic patches play a role in secondary nucleation. Single, double, and quadruple serine substitutions were made. Kinetic analyses of aggregation data at multiple monomer concentrations reveal that all seven mutants retain the dominance of secondary nucleation as the main mechanism of fibril proliferation. This finding highlights the generality of secondary nucleation and its independence of the detailed molecular structure. Cryo-electron micrographs reveal that the V18S substitution causes fibrils to adopt a distinct morphology with longer twist distance than variants lacking this substitution. Self- and cross-seeding data show that surface catalysis is only efficient between peptides of identical morphology, indicating a templating role of secondary nucleation with structural conversion at the fibril surface. Our findings thus provide clear evidence that the propagation of amyloid fibril strains is possible even in systems dominated by secondary nucleation rather than fragmentation.

Secondary Nucleation and the Conservation of Structural Characteristics of Amyloid Fibril Strains

“…In another recent study also involving the Aβ peptide, several sequence variants were investigated. It was shown that surface cross-nucleation was only efficient if the monomer had an innate preference (as judged from the fibrils formed de novo) for the structure of the seed fibril (Thacker et al, 2020). While these experiments are suggestive of some degree of specificity of fibril surface nucleation, they do not demonstrate the actual transmission of the template structure.…”

“…Cross-surface nucleation was found to be only efficient if monomer has same structural preference as seed fibril (Thacker et al, 2020).…”

“…While being strongly favored through binding to the surface of the seed fibrils, nucleation on fibril surfaces appears to still be subject to the influence of the solution conditions, as in the case of nucleation on foreign surfaces, such as the polymer-water or air-water interfaces (Bousset et al, 2013;Campioni et al, 2014). The only existing evidence for nucleation on fibril surfaces to be different from generic, non-specific surface nucleation is the data on the Aβ peptide, which suggests that this type of nucleation is inefficient if the monomer has a different structural preference from the seed fibril (Brännström et al, 2018;Thacker et al, 2020).…”

Illuminating amyloid fibrils: Fluorescence-based single-molecule approaches

“…Secondary nucleation is catalysed by the surface of pre-existing fibrils [48] , [49] , [50] , [51] , [52] . The properties of the ‘parent’ fibril that facilitate secondary nucleation remain to be established; however, it has been postulated that hydrophobic patches on the fibril surface [53] , and the ability of misfolded monomers to ‘recognize’ similar structures within the ‘parent’ fibril play a role in this process [54] . It was recently shown that decreasing the surface hydrophobicity of fibrils formed by the AD-associated Aβ(1-42) via serine substitution of hydrophobic residues exposed on the fibril surface, does not change the rate of secondary nucleation [53] .…”

“…The properties of the ‘parent’ fibril that facilitate secondary nucleation remain to be established; however, it has been postulated that hydrophobic patches on the fibril surface [53] , and the ability of misfolded monomers to ‘recognize’ similar structures within the ‘parent’ fibril play a role in this process [54] . It was recently shown that decreasing the surface hydrophobicity of fibrils formed by the AD-associated Aβ(1-42) via serine substitution of hydrophobic residues exposed on the fibril surface, does not change the rate of secondary nucleation [53] . However, both α-syn [55] and Aβ(1-42) [53] are able to ‘template’ secondary nuclei via the exposed surface residues of the mature fibrils, wherein monomers transiently bind and form oligomers with identical morphologies to those of the templating fibril.…”