Abstract
BackgroundIdentifying agents that inhibit amyloid beta peptide (Aβ) aggregation is the ultimate goal for slowing Alzheimer’s disease (AD) progression. This study investigated whether the glycoside asiaticoside inhibits Aβ1–42 fibrillation in vitro.MethodsFluorescence correlation spectroscopy (FCS), evaluating the Brownian diffusion times of moving particles in a small confocal volume at the single-molecule level, was used. If asiaticoside inhibits early Aβ1–42 fibrillation steps, more Aβs would remain free and rapidly diffuse in the confocal volume. In contrast, “weaker or no inhibition” permits a greater number of Aβs to polymerize into oligomers, leading to fibers and gives rise to slow diffusion times in the solution. Trace amounts of 5-carboxytetramethylrhodamine (TAMRA)-labeled Aβ1–42 in the presence of excess unlabeled Aβ1–42 (10 μM) was used as a fluorescent probe. Steady-state and kinetic-Thioflavin T (ThT) fluorospectroscopy, laser-scanning fluorescence microscopy (LSM), and transmission electron microscopy (TEM) were also used to monitor fibrillation. Binding of asiaticoside with Aβ1–42 at the atomic level was computationally examined using the Molegro Virtual Docker and PatchDock.ResultsWith 1 h of incubation time for aggregation, FCS data analysis revealed that the diffusion time of TAMRA-Aβ1–42 was 208 ± 4 μs, which decreased to 164 ± 8.0 μs in the presence of asiaticoside, clearly indicating that asiaticoside inhibited the early stages Aβ1–42 of fibrillation, leaving more free Aβs in the solution and permitting their rapid diffusion in the confocal volume. The inhibitory effects were also evidenced by reduced fiber formation as assessed by steady-state and kinetic ThT fluorospectroscopy, LSM, and TEM. Asiaticoside elongated the lag phase of Aβ1–42 fibrillation, indicating the formation of smaller amyloid species were impaired in the presence of asiaticoside. Molecular docking revealed that asiaticoside binds with amyloid intra- and inter-molecular amino acid residues, which are responsible for β-sheet formation and longitudinal extension of fibrils.ConclusionFinally, asiaticoside prevents amyloidogenesis that precedes neurodegeneration in patients with Alzheimer’s disease.
Highlights
Identifying agents that inhibit amyloid beta peptide (Aβ) aggregation is the ultimate goal for slowing Alzheimer’s disease (AD) progression
The autocorrelation function of TAMRAAβ1–42 in the (TAMRA-Aβ1–42 + unlabeled Aβ1–42) sample was best fitted with a one-component analysis model, which resulted in a diffusion time of 208 ± 4 μs (Figure 3A)
The diffusion time of TAMRAAβ1–42 in the (TAMRA-Aβ1–42 + unlabeled Aβ1–42 + 20 μM of asiaticoside) samples was reduced to 164 ± 8.0 μs (Figure 3B)
Summary
Identifying agents that inhibit amyloid beta peptide (Aβ) aggregation is the ultimate goal for slowing Alzheimer’s disease (AD) progression. Alzheimer’s disease (AD) is pathologically characterized by the fibrillar deposition of amyloid beta peptides (Aβ) in the brain [1]. Several key events are considered to be crucial, including the mechanism by which Aβ1–42 abandons its random coil-alpha helix conformation and adapts a β-sheet conformation, leading to oligomerization and matured fiber formation. These α-to-β conformational transitions are crucial for understanding the mechanisms of fiber formation and inhibition by agents capable of inhibiting the fibrillation process
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