Reversible Mechanical Unzipping of Amyloid β-Fibrils

Abstract

Amyloid fibrils are self-associating filamentous structures, the deposition of which is considered to be one of the most important factors in the pathogenesis of Alzheimer's disease and various other disorders. Here we used single molecule manipulation methods to explore the mechanics and structural dynamics of amyloid fibrils. In mechanically manipulated amyloid fibrils, formed from either amyloid ␤ (A␤) peptides 1-40 or 25-35, ␤-sheets behave as elastic structures that can be "unzipped" from the fibril with constant forces. The unzipping forces were different for A␤1-40 and A␤25-35. Unzipping was fully reversible across a wide range of stretch rates provided that coupling, via the ␤-sheet, between bound and dissociated states was maintained. The rapid, cooperative zipping together of ␤-sheets could be an important mechanism behind the self-assembly of amyloid fibrils. The repetitive force patterns contribute to a mechanical fingerprint that could be utilized in the characterization of different amyloid fibrils.Amyloid fibrils are self-associating filamentous structures formed from the 39 -43-residue-long amyloid ␤-peptide (A␤) 1 or its subfragments (1). The deposition of amyloid oligomers (2) and fibrils is considered to be one of the most important factors in the pathogenesis of Alzheimer's disease (3) and other disorders (4). The structure of A␤-fibrils has for long been enigmatic because of insoluble aggregate formation that precludes the use of standard structural methods such as x-ray crystallography and solution NMR. Recent data from site-directed spin labeling (5), and particularly from solid-state NMR experiments (6, 7), have formed the basis of a high resolution model of the A␤1-40 fibril: ␤-hairpins lying perpendicular to the fibril axis are associated into ␤-sheets that line up to form protofilaments, which are then assembled parallel into fibrils. Protofilaments are thus thought to represent an ϳ2-3-nm-diameter structural unit within the amyloid fibril (1). During amyloidogenesis the formation of fibrils is preceded by the appearance of globular aggregates that are thought to fuse, by not fully understood mechanisms, into fibrillar structures (8). Recently, curved, beaded, ϳ200-nm-long and ϳ6 -8-nm-wide fibrillar precursors were described to appear in the amyloidogenetic pathway, which were called protofibrils (9 -12). The protofibrils are thought to go through a structural transition on their way to forming the amyloid fibril. The exact nature of structural dynamics within the amyloid fibril related to amyloidogenesis, however, remains to be resolved.Single molecule manipulation experiments have in the recent past provided unique and unprecedented insights not only into the structure and elasticity but also into mechanically driven transitions of molecular systems (13)(14)(15)(16)(17)(18)(19)(20)(21). In the present work we mechanically manipulated amyloid fibrils formed from either A␤1-40 or A␤25-35 peptides. We showed that filamentous entities most likely corresponding to ␤-sheets can be "unzipp...

Nanomechanics of Amyloid Materials Studied by Atomic Force Microscopy

“…This method avoids indentation into the matrix, thus reduces the possibility of picking up multiple molecules in a single pull. Fitting the force curve with WLC model gave a persistence length of 0.35 ± 0.05 nm, which agrees pretty well with previous results on amyloid fibrils (Kellermayer, et al, 2005;. In addition, FT-IR spectroscopy and dye (Congo red and thioflavin T) staining confirmed the presence of -sheet and amyloid structures in the sample.…”

“…All of the force curves exhibited non-linear elastic mechanical response followed by one or several rupture events. The absence of force plateau, which is a characteristic pattern of the unzipping of -sheets from the fibril, implies that the high packing density in mature glucagon amyloid fibrils allows a stable confinement of the peptide molecules in the twisted fibrillar structure, in contrast to A amyloid fibrils (Karsai et al, 2006;Kellermayer, et al, 2005). The stable structure is further confirmed by repeatedly stretching a single fibril for 1000 times.…”

“…This implies that the associated state of thesheets is highly favored and stabilized within the amyloid fibrils (Kellermayer, et al, 2005), which partly explains the high stability of amyloid fibrils. (Kellermayer, et al, 2005).…”

“…In the AFM manipulation of the A 1-40 and A 25-35 fibrils (Kellermayer et al, 2005), the resulting force curves exhibited two distinct mechanical responses: (1) fully reversible, non-linear elasticity with WLC fitted contour length often exceeding 100 nm ( Figure 6A); (2) force platform characterized with long distance constant force stretching followed by abrupt force drop ( Figure 6B). In most cases, the two patterns are superposed onto each other ( Figure 6C, 6D).…”

Single-molecule force spectroscopy: optical tweezers, magnetic tweezers and atomic force microscopy

“…More recently, Lim and coworkers showed that the elastic properties of blood clots are mainly determined by the coiled-coil helices of fibrinogen 142 . In another example, Kellermayer and coworkers described the structure and self assembly mechanism of amyloid β-fibrils 143 and myosin thick filaments 144 .…”