Quantification of amyloid fibril polymorphism by nano-morphometry reveals the individuality of filament assembly

Abstract

Amyloid fibrils are highly polymorphic structures formed by many different proteins. They provide biological function but also abnormally accumulate in numerous human diseases. The physicochemical principles of amyloid polymorphism are not understood due to lack of structural insights at the single-fibril level. To identify and classify different fibril polymorphs and to quantify the level of heterogeneity is essential to decipher the precise links between amyloid structures and their functional and disease associated properties such as toxicity, strains, propagation and spreading. Employing gentle, force-curve based AFM, we produce detailed images, from which the 3D reconstruction of individual filaments in heterogeneous amyloid samples is achieved. Distinctive fibril polymorphs are then classified by hierarchical clustering, and sample heterogeneity is objectively quantified. These data demonstrate the polymorphic nature of fibril populations, provide important information regarding the energy landscape of amyloid self-assembly, and offer quantitative insights into the structural basis of polymorphism in amyloid populations.

Three-dimensional reconstruction of individual helical nano-filament structures from atomic force microscopy topographs

“…The deconvolution algorithm and resulting lateral sampling enhancement are demonstrated on an AFM topograph of an amyloid fibril formed from a short amyloid forming peptide with the amino acid sequence HYFNIF (Materials and Methods and [17]. The fibril example is first traced from the AFM image across the fibril central line and subsequently straightened and interpolated to an evenly spaced pixel grid [26], while maintaining pixel size identical to that in raw data.…”

“…3D surface envelope reconstruction of amyloid fibrils from AFM topography. (a) AFM topology image of amyloid fibrils assembled from short peptide with the sequence HYFNIF [17,24]. The scale bars represent 1 μm.…”

“…Furthermore, specific structural polymorphs assembled from the same amyloid forming protein could be patient- or disease-specific and may correlate with neurodegenerative disease aetiology [14]. Due to the unresolved nature of the amyloid assembly structure-function relationship, fibrillar amyloid specimen are widely studied using AFM imaging to resolve the mechanistic roles of amyloid assembly and polymorphism [17].…”

Three-dimensional reconstruction of individual helical nano-filament structures from atomic force microscopy topographs

“…The deconvolution algorithm and resulting lateral sampling enhancement are demonstrated on an AFM topograph of amyloid fibrils formed from a short amyloid forming peptide with the amino acid sequence HYFNIF (Materials and Methods and [17]). The fibril example is first traced from the AFM image across the fibril central line and subsequently straightened and interpolated to an evenly spaced pixel grid [26], while maintaining pixel size identical to that in raw data.…”

“…Furthermore, specific structural polymorphs assembled from the same amyloid forming protein could be patient-or disease-specific and may correlate with neurodegenerative disease aetiology [14]. Due to the unresolved nature of the amyloid assembly structure-function relationship, fibrillar amyloid specimens are widely studied using AFM imaging to resolve the mechanistic roles of amyloid assembly and polymorphism [17].…”