Probing water-accessibility in HET-s(218-289) amyloid fibrils by solid-state NMR.
 

Probing water-accessibility in HET-s(218-289) amyloid fibrils by solid-state NMR.

Probing water-accessibility in HET-s(218-289) amyloid fibrils by solid-state NMR.

J Mol Biol. 2010 Nov 18;

Authors: Van Melckebeke H, Schanda P, Gath J, Wasmer C, Verel R, Lange A, Meier BH, Böckmann A

Despite its importance in the context of conformational diseases, structural information is still sparse for protein fibrils. Hydrogen/deuterium exchange measurements of backbone amides allow to identify hydrogen-bonding patterns and reveal pertinent information about the amyloid ?-sheet architecture. However, they provide only little information about the identity of residues exposed to solvent or which are buried inside the fibril core. NMR spectroscopy is a potent method to identify solvent accessible residues in proteins via the observation of polarization transfer between chemically exchanging sidechain protons and water protons. We show here that the combined use of highly deuterated samples and fast magic-angle spinning (MAS) greatly attenuates unwanted spin diffusion and allows identifying 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 measurements carried out at physiological pH, where the protofibrils assemble together into thicker fibrils and demonstrate that each protofibril of HET-s(218-289) is surrounded by water, excluding extended dry inter-fibril 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 detailed structural features of amyloid fibrils in general.

PMID: 21094164 [PubMed - as supplied by publisher]



Source: PubMed