Oral Presentation The 46th Lorne Conference on Protein Structure and Function 2021

Fluorescent probes for super-resolution imaging of protein aggregates (#28)

Amandeep Kaur 1 2 , Margie Sunde 1 2 , Elizabeth New 2 3
  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

Molecular interactions at the nanometre-level critically influence the aggregation of misfolded proteins into macromolecular structures called amyloids that are hallmarks of many neurodegenerative diseases.1 In Alzheimer’s disease (AD) and frontotemporal lobar degeneration (FTLD), the amyloid-β, tau and TDP-43 proteins can be found assembled in amyloid fibrils, paired helical filaments or other aggregates where the morphology is less defined.2 A key question in this field is the nature of the differences between globular and fibrillar aggregates of these proteins.3,4 Standard optical techniques are limited by optical diffraction, examine ensemble behaviour and provide no insight into the structural details governing protein aggregation and its impact on other cellular factors.5,6 Therefore, the ability to visualise molecular-level structure and organisation of proteins in these diseases is central to gain thorough understanding of the mechanisms of protein aggregation in neurodegeneration.

In recent years, super-resolution microscopy has revolutionised the study of biological and synthetic nanostructures by breaking the diffraction limit, and allowing visualisation of cells and materials on the molecular scale.7 While this Nobel Prize-winning technology demonstrates great potential for biomedical researchers wishing to unravel molecular-level interactions, the quantitative information obtainable and the progress in this field, is greatly limited by the limited number and poor photophysical properties of existing fluorophores.6

We have developed a novel coumarin-based fluorescent amyloid sensor (AmyBlink-1) that utilises the amyloidophilic properties of the benzothiazole scaffold. Upon interaction with amyloid fibrils, AmyBlink-1 exhibits a dramatic change in its emission profile: turn-on in fluorescence and blue-shifted emission maximum. In addition, the sensor undergoes photoswitching properties allowing for super-resolution imaging of amyloid fibrils. This type of sensor will be a valuable tool for researchers working towards understanding neurodegenerative pathologies.

  1. Benilova, I.; Karran, E.; De Strooper, B. The Toxic Aβ Oligomer and Alzheimer’s Disease: An Emperor in Need of Clothes. Nat. Neurosci. 2012, 15 (3), 349–357. https://doi.org/10.1038/nn.3028.
  2. Iadanza, M. G.; Jackson, M. P.; Hewitt, E. W.; Ranson, N. A.; Radford, S. E. A New Era for Understanding Amyloid Structures and Disease. Nat. Rev. Mol. Cell Biol. 2018, 19 (12), 755–773. https://doi.org/10.1038/s41580-018-0060-8.
  3. Fodero-Tavoletti, M. T.; Villemagne, V. L.; Rowe, C. C.; Masters, C. L.; Barnham, K. J.; Cappai, R. Amyloid-β: The Seeds of Darkness. Int. J. Biochem. Cell Biol. 2011, 43 (9), 1247–1251. https://doi.org/https://doi.org/10.1016/j.biocel.2011.05.001.
  4. Larson, M. E.; Lesné, S. E. Soluble Aβ Oligomer Production and Toxicity. J. Neurochem. 2012, 120 (s1), 125–139. https://doi.org/10.1111/j.1471-4159.2011.07478.x.
  5. Rust, M. J.; Bates, M.; Zhuang, X. Sub-Diffraction-Limit Imaging by Stochastic Optical Reconstruction Microscopy (STORM). Nat. Methods 2006, 3 (10), 793–795. https://doi.org/10.1038/nmeth929.
  6. Kaur, A.; New, E. J.; Sunde, M. Strategies for the Molecular Imaging of Amyloid and the Value of a Multimodal Approach. ACS Sensors 2020, 5 (8), 2268–2282. https://doi.org/10.1021/acssensors.0c01101.
  7. Schermelleh, L., Ferrand, A., Huser, T. et al. Super-resolution microscopy demystified. Nat Cell Biol 21, 72–84 (2019). https://doi.org/10.1038/s41556-018-0251-8