Biophysical studies of the development of amyloid fibrils from a peptide fragment of cold shock protein B

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Abstract
The peptide CspB-1, which represents residues 1–22 of the cold shock protein CspB from Bacillus subtilis, has been shown to form amyloid fibrils when solutions containing this peptide in aqueous (50%) acetonitrile are diluted in water [M. Großet al. (1999) Protein Science8, 1350–1357] We established conditions in which reproducible kinetic steps associated with the formation of these fibrils can be observed. Studies combining these conditions with a range of biophysical methods reveal that a variety of distinct events occurs during the process that results in amyloid fibrils. A CD spectrum indicative of β structure is observed within 1 min of the solvent shift, and its intensity increases on a longer timescale in at least two kinetic phases. The characteristic wavelength shift of the amyloid-binding dye Congo Red is established within 30 min of the initiation of the aggregation process and corresponds to one of the phases observed by CD and to changes in the Fourier transform-infrared spectrum indicative of β structure. Short fibrillar structures begin to be visible under the electron microscope after these events, and longer, well-defined amyloid fibrils are established on a timescale of hours. NMR spectroscopy shows that there are no significant changes in the concentration of monomeric species in solution during the events leading to fibril formation, but that soluble aggregates too large to be visible in NMR spectra are present throughout the process. A model for amyloid formation by this peptide is presented which is consistent with these kinetic data and with published work on a variety of disease-related systems. These findings support the concept that the ability to form amyloid fibrils is a generic property of polypeptide chains, and that the mechanism of their formation is similar for different peptides and proteins.