Monitoring amyloid formation from natively folded proteins to amyloid fibrils by NMR spectroscopy.

Amyloids are insoluble protein aggregates that arise through protein misfolding and are associated with numerous neurodegenerative disorders, systemic amyloidosis, and other human diseases. Nuclear magnetic resonance (NMR) spectroscopy, encompassing both solution-state and solid-state approaches, offers unique capabilities to investigate each stage of the complex amyloid formation pathway, from soluble monomeric precursors through transient intermediate states and oligomeric assemblies to highly ordered mature fibrils. This residue-specific and atomic-level characterization of the aggregation process is difficult to achieve using other biophysical techniques. NMR is particularly well suited for elucidating the mechanisms of amyloid formation because it provides detailed structural and dynamic information across multiple aggregation states at molecular resolution. This review highlights the application of solution-state and solid-state NMR methods in characterizing the conversion of natively folded amyloidogenic proteins into amyloid fibrils, with particular emphasis on the identification and characterization of transient intermediates, including both observable and "invisible" species, high-molecular-weight oligomers, and insoluble fibrillar aggregates. Representative examples of natively folded amyloidogenic precursor proteins are discussed, together with an overview of the NMR method used to elucidate their misfolding and aggregation mechanisms. Collectively, these studies demonstrate the power of NMR spectroscopy in providing mechanistic and structural insights into amyloid formation at residue-level resolution.