Template-Assisted Lateral Growth of Amyloid-β42 Fibrils Studied by Differential Labeling with Gold Nanoparticles

Amyloid-β protein (Aβ) aggregation into amyloid fibrils is central to the origin and development of Alzheimer’s disease (AD), yet this highly complex process is poorly understood at the molecular level. Extensive studies have shown that Aβ fibril growth occurs through fibril elongation, whereby soluble molecules add to the fibril ends. Nevertheless, fibril morphology strongly depends on aggregation conditions. For example, at high ionic strength, Aβ fibrils laterally associate into bundles. To further study the mechanisms leading to fibril growth, we developed a single-fibril growth assay based on differential labeling of two Aβ42 variants with gold nanoparticles. We used this assay to study Aβ42 fibril growth under different conditions and observed that bundle formation is preceded by lateral interaction of soluble Aβ42 molecules with pre-existing fibrils. Based on this data, we propose template-assisted lateral fibril growth as an additional mechanism to elongation for Aβ42 fibril growth

Amount of DKP, diclofenac, DKP-diclofenac conjugate, TAT-diclofenac conjugate and TAT in the skin after permeation experiments (donor amount of 150 μg).

<p>Data is represented as mean ± sem (n = 4), * P<0.05.</p

Human skin permeation of diclofenac alone, diclofenac as admixture with DKP, diclofenac as DKP conjugate and diclofenac as TAT conjugate.

<p>The data is represented as mean ± sem (n = 5), *P<0.05.</p

Representation of the linker system used in the (a) <i>C</i>-terminal analogs (b) <i>N</i>-terminal analogs.

<p>Representation of the linker system used in the (a) <i>C</i>-terminal analogs (b) <i>N</i>-terminal analogs.</p

RP-HPLC Chromatogram of the enzymatic evaluation of the model substrate.

<p>Conditions: lineal gradient of 0–100% MeCN (0.036% TFA) in H₂O (0.045% TFA) in 15 min. Flux of 1 mL/min, in a symmetry column C₁₈ (4.6 mm×150 mm), Waters, 100 Å, 5 µm, detection at 220 nm.</p

Summary of permeation data for diclofenac and peptide shuttles.

<p>Summary of permeation data for diclofenac and peptide shuttles.</p

Representation of the peptide bound to PEGA₁₉₀₀ resin using the PLL linker (drawn in green).

<p>Representation of the peptide bound to PEGA₁₉₀₀ resin using the PLL linker (drawn in green).</p

Representation of (a) the model substrate, (b) the <i>C</i>-terminal-anchored analogs and control peptides, (c) the <i>N</i>-terminal-anchored analogs and control peptides.

<p>Representation of (a) the model substrate, (b) the <i>C</i>-terminal-anchored analogs and control peptides, (c) the <i>N</i>-terminal-anchored analogs and control peptides.</p

Representation of the peptides used to confirm the esterase activity and capacity to process on solid-phase.

<p>(a) <i>C</i>-terminal-anchored analogs directly bounded on solid-phase. (b) <i>C</i>-terminal control peptides for the fluorescence assay directly bounded on solid-phase.</p

Characteristics and performance of the tested algorithms.

<p>The scoring function in the deterministic algorithms is based on achieving zero signal overlap (<i>Scoring = Ni</i>, where N_i: number of fragments in the mixture) while in stochastic algorithms the scoring function is based on achieving minimal signal overlap (, where N_ov: number of overlapped signals of compound <i>i</i>, and N_t: total number of signals of compound <i>i</i>).</p