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Updated: Sep 28, 2026
Biochemical Purification and Proteomic Characterization of Amyloid Fibril Cores from the Brain
Published on: April 28, 2022
AA protein in experimental murine AA amyloid fibrils: a high resolution ultrastructural and immunohistochemical study
Sadayuk Inoue1, Mie Kuroiwa, Robert Kisilevsky
1Department of Anatomy and Cell Biology, McGill University, Montreal, Quebec, Canada. sadayuki.inoue@mcgill.ca
Abstract:
In a previous study, the fibrils of experimental murine AA amyloid were found to be microfibril-like structures with the AA protein (in the form of 1 nm wide flexible filaments) on their exterior surface. In this study, we have re-examined the AA amyloid fibrils with advanced methods of cryofixation and freeze substitution which are known to retain ultrastructural detail as close as possible to the living state. The observations were compared to those obtained with conventional methods of aldehyde fixation. Cryofixation and freeze substitution confirmed the microfibril-like nature of the inner part of the AA amyloid fibril. The AA protein was present on the exterior surface in the form of 3 nm wide 'helical rods' formed by the tight coiling of the 1 nm wide AA protein flaments. The 'helical rods' were arranged parallel to the axis of the fibril and to one another with a uniform center-to-center distance of 5 nm. This arrangement was fully preserved in amyloid fibrils after cryofixation and freeze substitution, but was present in only some areas of formaldehyde fixed mouse spleen AA amyloid This conformation and orientation of AA protein is likely to be that in its native state, given the ability of these advanced methods of biological preservation to retain structures close to that of the living state. This information shoulld be of considerable value in comparing the structure of amyloid fibrils observed in situ with those isolated from tissue or generated in vitro.
Insights
Advanced cryofixation reveals amyloid AA protein forms helical rods on fibril exteriors. This structure, preserved by cryofixation, likely represents the native state of amyloid AA protein.
Area of Science:
- Biochemistry
- Cell Biology
- Structural Biology
Background:
- Previous studies identified AA amyloid fibrils with AA protein on their exterior.
- The exact ultrastructure and native conformation of AA protein in amyloid fibrils remained unclear.
Purpose of the Study:
- To re-examine the ultrastructure of experimental murine AA amyloid fibrils using advanced preservation techniques.
- To compare findings from cryofixation and freeze substitution with conventional aldehyde fixation methods.
- To elucidate the native conformation and arrangement of AA protein on amyloid fibrils.
Main Methods:
- Utilized cryofixation and freeze substitution for high-fidelity ultrastructural preservation.
- Employed conventional aldehyde fixation for comparative analysis.
- Examined experimental murine AA amyloid fibrils.
Main Results:
- Cryofixation confirmed the microfibrillar core of AA amyloid fibrils.
- AA protein was observed on the exterior as 3 nm wide 'helical rods', formed by coiled 1 nm filaments.
- These helical rods exhibited parallel arrangement with 5 nm center-to-center spacing, a structure well-preserved by cryofixation but inconsistently by aldehyde fixation.
Conclusions:
- The observed helical rod structure of AA protein on amyloid fibrils likely represents its native state due to advanced preservation methods.
- Findings provide valuable insights for comparing in situ amyloid structures with isolated or in vitro generated fibrils.
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