Abstract
Amyloid β proteins spontaneously form fibrils in vitro that vary in their thermodynamic stability and in morphological characteristics such as length, width, shape, longitudinal twist, and the number of component filaments. It is vitally important to determine which variant best represents the type of fibril that accumulates in Alzheimer disease. In the present study, the nature of morphological variation was examined by dark-field and transmission electron microscopy in a preparation of seeded amyloid β protein fibrils that formed at relatively low protein concentrations and exhibited remarkably high thermodynamic stability. The number of filaments comprising these fibrils changed frequently from two to six along their length, and these changes only became apparent when mass-per-length (MPL) determinations are made with sufficient resolution. The MPL results could be reproduced by a simple stochastic model with a single adjustable parameter. The presence of more than two primary filaments could not be discerned by transmission electron microscopy, and they had no apparent relationship to the longitudinal twist of the fibrils. However, the pitch of the twist was strongly affected by the pH of the negative stain. We conclude that highly stable amyloid fibrils may form in which a surprising amount of intrinsic linear heterogeneity may be obscured by MPL measurements of insufficient resolution, and by the negative stains used for imaging fibrils by electron microscopy.
Highlights
Folded within fibrils, yet the models emerging from these efforts differ significantly [1,2,3,4,5,6,7,8]
To understand how fibril morphology depends on the manner in which their morphologies are characterized
We have found that the morphology of these highly stable fibrils was strongly influenced by the pH of negative stains
Summary
Folded within fibrils, yet the models emerging from these efforts differ significantly [1,2,3,4,5,6,7,8]. To quantify the distribution of internodal distances and determine whether there were subpopulations of fibrils with different morphologies, histograms of the observed internodal distances were generated for each stain/pH combination (Fig. 5). For 873 measurements derived from 40 ϫ 80 nm windows, there are 5 distinct peaks corresponding to the MPL values expected for 2, 3, 4, 5, and 6 filaments per fibril.
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