Oral Presentation 14th Australian Peptide Conference 2022

Fluorescent sensors for super-resolution imaging of amyloid fibril assembly (#42)

Amandeep Kaur 1 2 , Kai Kikuchi 1 2 , Liam D Adair 3 , Elizabeth New 2 3 , Margie Sunde 1 2
  1. School of Medical Sciences, The University of Sydney, Sydney, New South Wales, Australia
  2. Sydney Nano, The University of Sydney, Sydney, New South Wales, Australia
  3. School of Chemistry, The University of Sydney, Sydney, New South Wales, Australia

Many soluble proteins can self-assemble into macromolecular structures called amyloids, a subset of which are implicated in a range of neurodegenerative disorders. A key question in this field is the nature of the differences between globular and fibrillar aggregates of these proteins and how the histopathologic patterns relate to the different types of neurodegenerative conditions. Therefore, an ability to visualise the molecular-level organisation, structure and distribution of proteins in these proteinopathies is critical to gain a thorough understanding of the mechanisms underlying the associated neurodegeneration.  The nanoscale size and structural heterogeneity of prefibrillar and early aggregates, as well as mature amyloid fibrils, pose significant challenges for the quantification of amyloid morphologies. In this presentation, I will describe our work on development of a fluorescent amyloid sensor AmyBlink-1 and its application in super-resolution imaging of amyloid structures. AmyBlink-1 exhibits a 5-fold increase in ratio of the green (thioflavin T) to red (Alexa Fluor 647) emission intensities upon interaction with amyloid fibrils. Using AmyBlink-1, we performed nanoscale imaging of four different types of amyloid fibrils, achieving a resolution of ≈30 nm. AmyBlink-1 enables nanoscale visualization and subsequent quantification of morphological features, such as the length and skew of individual amyloid aggregates formed at different times along the amyloid assembly pathway. Quantitative analysis of the heterogeneity, on the nanoscale, during the formation and deposition of fibrils will open new doors to understanding the molecular mechanisms of amyloid-related pathologies.

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