Chemical dissection and reassembly of amyloid fibrils formed by a peptide fragment of transthyretin

Abstract

We have examined the chemical dissection and subsequent reassembly of fibrils formed by a ten-residue peptide to probe the forces that drive the formation of amyloid. The peptide, TTR10-19, encompasses the A strand of the inner β-sheet structure that lines the thyroid hormone binding site of the human plasma protein transthyretin. When dissolved in water under low pH conditions the peptide readily forms amyloid fibrils. Electron microscopy of these fibrils indicates the presence of long (>1000 nm) rigid structures of uniform diameter (approximately 14 nm). Addition of urea (3 M) to preformed fibrils disrupts these rigid structures. The partially disrupted fibrils form flexible ribbon-like arrays, which are composed of a number of clearly visible protofilaments (3-4 nm diameter). These protofilaments are highly stable, and resist denaturation in 6 M urea at 75 °C over a period of hours. High concentrations (>50 %, v/v) of 2,2,2-trifluoroethanol also dissociate TTR10-19 fibrils to the constituent protofilaments, but these slowly dissociate to monomeric, soluble peptides with extensive α-helical structure. Dilution of the denaturant or co-solvent at the stage when dissociation to protofilaments has occurred results in the efficient reassembly of fibrils. These results indicate that assembly of fibrils from protofilaments involves relatively weak and predominantly hydrophobic interactions, whereas assembly of peptides into protofilaments involves both electrostatic and hydrophobic forces, resulting in a highly stable and compact structures.