Despite the importance of protein fibrils in the context of conformational diseases, information on their structure is still sparse. Hydrogen/deuterium exchange measurements of backbone amide protons allow the identification hydrogen-bonding patterns and reveal pertinent information on the amyloid β-sheet architecture. However, they provide only little information on the identity of residues exposed to solvent or buried inside the fibril core. NMR spectroscopy is a potent method for identifying solvent-accessible residues in proteins via observation of polarization transfer between chemically exchanging side-chain protons and water protons. We show here that the combined use of highly deuterated samples and fast magic-angle spinning greatly attenuates unwanted spin diffusion and allows identification of polarization exchange with the solvent in a site-specific manner. We apply this measurement protocol to HET-s(218-289) prion fibrils under different conditions (including physiological pH, where protofibrils assemble together into thicker fibrils) and demonstrate that each protofibril of HET-s(218-289), is surrounded by water, thus excluding the existence of extended dry interfibril contacts. We also show that exchangeable side-chain protons inside the hydrophobic core of HET-s(218-289) do not exchange over time intervals of weeks to months. The experiments proposed in this study can provide insight into the detailed structural features of amyloid fibrils in general.
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