Fine structure study of Aβ_1–42fibrillogenesis with atomic force microscopy

Abstract

One of the hallmarks of Alzheimer's disease is the self-aggregation of the amyloid beta peptide (Abeta) in extracellular amyloid fibrils. Among the different forms of Abeta, the 42-residue fragment (Abeta1-42) readily self-associates and forms nucleation centers from where fibrils can quickly grow. The strong tendency of Abeta1-42 to aggregate is one of the reasons for the scarcity of data on its fibril formation process. We have used atomic force microscopy (AFM) to visualize in liquid environment the fibrillogenesis of synthetic Abeta1-42 on hydrophilic and hydrophobic surfaces. The results presented provide nanometric resolution of the main structures characteristic of the several steps from monomeric Abeta1-42 to mature fibrils in vitro. Oligomeric globular aggregates of Abeta1-42 precede the appearance of protofibrils, the first fibrillar species, although we have not obtained direct evidence of oligomer-protofibril interconversion. The protofibril dimensions deduced from our AFM images are consistent with a model that postulates the stacking of the peptide in a hairpin conformation perpendicular to the long axis of the protofibril, forming single beta-sheets ribbon-shaped. The most abundant form of Abeta1-42 fibril exhibits a nodular structure with a ~100-nm periodicity. This length is very similar 1) to the length of protofibril bundles that are the dominant feature at earlier stages in the aggregation process, 2) to the period of helical structures that have been observed in the core of fibrils, and 3) to the distance between regularly spaced, structurally weak fibril points. Taken together, these data are consistent with the existence of a ~100-nm long basic protofibril unit that is a key fibril building block.

Hsp104 Targets Multiple Intermediates on the Amyloid Pathway and Suppresses the Seeding Capacity of Aβ Fibrils and Protofibrils

“…24 Protofibrils are on-pathway intermediates and convert into mature amyloid fibrils. 26,[29][30][31] To probe the ability of Hsp104 to interact with these structures and to determine the consequences of such interactions on the stability and fibrillization of protofibrils, freshly isolated protofibrils (Fig. 3a) were co-incubated with Hsp104 and the aggregation was monitored by ThT fluorescence, TEM, and SDS-PAGE.…”

A truncated and dimeric format of an Affibody library on bacteria enables FACS‐mediated isolation of amyloid‐beta aggregation inhibitors with subnanomolar affinity

“…According to the amyloid cascade hypothesis, the amyloid beta (Ab) peptide is central to the pathogenesis of AD [4]. It has been hypothesized that the aggregation process of these peptides (mainly 40 and 42 amino acids in length) into oligomers, protofibrils and fibrils play a pivotal role in the neuropathology of the disorder [5,6]. Consequently, therapeutic interventions targeting the production and aggregation, as well as clearance of Ab from the brain are under investigation [2,7,8].…”

Direct Observation of Amyloid Nucleation under Nanomechanical Stretching

“…Background appears to be flat, however, force spectroscopy on the background reveals that the background consists of enriched SELP layer that even some SELP fold and form fiber, this layer didn't shows any depletion region. [19] Some profiles showed a single large step increase which was quantized at 2.1±0.1 nm (n=7) and 3.5±0.2 nm (n=32), indicating that multiple layers of beta sheet were formed instantly. The fully grown height of the nucleus coincides with the height of preformed SELP which is 3.7±0.2 nm (n=7).…”

“…In addition to studies in bulk solution, amyloid formation on surfaces has been actively investigated since it may more closely mimic physiological environments such as cell membrane [13]. Surface accelerated assembly kinetics as well as directed growth of amyloid on surfaces has been reported using various peptides [14][15][16][17][18][19].…”

“…When a sharpened tip was used to minimize the effect of tip broadening, the width of the mature fiber was measured to be 13±4 nm (n=14) which is similar to the width of other amyloids. [19] Assuming that the silk unit (GAGAGS) 8 in SELP drives amyloid formation and one silk unit participates in one beta hairpin structure, the approximate width of amyloid which is determined by the half number of silk units is calculated to be ~8.6 nm (24 residues  0.36 nm/residue). This is very close to the width measured using the sharpened tip and suggests that the silk unit plays a key role in amyloid assembly.…”