Cryo-EM of amyloid fibrils: Advances, challenges, and translational applications

Cryo-electron microscopy (cryo-EM) has emerged as a transformative technique in structural biology, driving an exponential increase in the number of amyloid fibril structures deposited in the Protein Data Bank. Current workflows now routinely achieve the near-atomic resolution essential for understanding the molecular pathology of neurodegenerative and metabolic disorders. This chapter provides a comprehensive overview of the field, tracing the evolution from early structural studies to modern pipelines powered by deep-learning algorithms for helical reconstruction. We address the persistent bottleneck of sample preparation, detailing the physical challenges of fibril clumping and interfacial adsorption, alongside emerging solutions to these problems. Furthermore, we examine the complexities of structural polymorphism, discussing how environmental factors and seeding drive conformational diversity and how this heterogeneity impacts biochemical assays. Finally, we highlight the translational value of these high-resolution maps, demonstrating how they enable the rational, structure-based design of peptide inhibitors and small molecules capable of disaggregating pathogenic fibrils.