title: Amide proton solvent protection in amylin fibrils probed by quenched hydrogen exchange NMR.
abstract: Amylin is an endocrine hormone that accumulates in amyloid plaques in patients with advanced type 2 diabetes. The amyloid plaques have been implicated in the destruction of pancreatic β-cells, which synthesize amylin and insulin. To better characterize the secondary structure of amylin in amyloid fibrils we assigned the NMR spectrum of the unfolded state in 95% DMSO and used a quenched hydrogen-deuterium exchange technique to look at amide proton solvent protection in the fibrils. In this technique, partially exchanged fibrils are dissolved in 95% DMSO and information about amide proton occupancy in the fibrils is determined from DMSO-denatured monomers. Hydrogen exchange lifetimes at pH 7.6 and 37°C vary between ∼5 h for the unstructured N-terminus to 600 h for amide protons in the two β-strands that form inter-molecular hydrogen bonds between amylin monomers along the length of the fibril. Based on the protection data we conclude that residues A8-H18 and I26-Y37 comprise the two β-strands in amylin fibrils. There is variation in protection within the β-strands, particularly for strand β1 where only residues F15-H18 are strongly protected. Differences in protection appear to be due to restrictions on backbone dynamics imposed by the packing of two-layers of C2-symmetry-related β-hairpins in the protofilament structure, with strand β1 positioned on the surface and β2 in the interior.
authors: Department of Molecular and Cell Biology, University of Connecticut, Storrs, Connecticut, United States of America. andrei@uconn.edu
authors: Andrei T
authors: AT
authors: Alexandrescu
mesh_terms: D000577:Amides / Q000737:chemistry*; D006801:Humans; D058228:Islet Amyloid Polypeptide / Q000737:chemistry*; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D056004:Molecular Dynamics Simulation; D055503:Protein Multimerization; D017433:Protein Structure, Secondary; D011522:Protons; D012997:Solvents / Q000737:chemistry*
chemical_list: D000577:Amides; D058228:Islet Amyloid Polypeptide; D011522:Protons; D012997:Solvents