2.19 Biophysical analysis of amyloid formation

Amyloid formation is a fascinating process with both biomedical and materials science relevance. Amyloids can be pathological, but also possess interesting potential for use as nanobiomaterials. Although amyloids have long been the focus of intense study, the amyloid formation mechanism stays unclear, including the factors that initiate and drive the aggregation process. This chapter describes the application of advanced biophysical methods, such as (single-molecule) fluorescence spectroscopy, electron paramagnetic resonance (EPR) spectroscopy, circular dichroism (CD) spectroscopy, and high-resolution atomic force microscopy (AFM), for exploration of amyloid characteristics. Collectively, the selection of techniques provides access to all stages of amyloid aggregation: from monomers to oligomeric intermediates, and mature fibrils. Single-molecule fluorescence is shown to provide information on monomer structure and flexibility. On an ensemble level, conformational information is acquired using CD spectroscopy and EPR spectroscopy. High-resolution AFM enables detailed exploration of morphological and mechanical properties of fibrillar structures. The biophysical research approaches described are widely applicable to the broader genre of amyloid proteins to uncover the mysteries that underlie the complex biophysics of amyloid formation and their optimal utilization in bionanotechnology