Evidence for a mechanism of amyloid formation involving molecular reorganisation within native-like precursor aggregates

Year: 2005

Authors: Plakoutsi G., Bemporad F., Calamai M., Taddei N., Dobson CM., Chiti F.

Autors Affiliation: Dipartimento di Scienze Biochimiche, Universita` di Firenze, Viale Morgagni 50 50134 Firenze, Italy;
Department of Chemistry University of Cambridge Lensfield Road, Cambridge CB2 1EW, UK

Abstract: The aggregation of the alpha/beta protein acylphosphatase from Sulfolobus solfataricus has been studied under conditions in which the protein maintains a native-like, although destabilised, conformation and that therefore bear resemblance to a physiological medium. Static and dynamic light-scattering measurements indicate that under these conditions the protein aggregates rapidly, within two minutes. The initial aggregates are enzymatically active and have a secondary structure that is not yet characterized by the high content of cross-beta structure typical of amyloid, as inferred from Fourier transform infra-red and circular dichroism measurements. These species then convert slowly into enzymatically inactive aggregates that bind thioflavin T and Congo red, characteristic of amyloid structures, and contain extensive beta-sheet structure. Transmission electron microscopy reveals the presence in the latter aggregates of spherical species and thin, elongated protofibrils, both with diameters of 3-5 nm. Kinetic tests reveal that this process occurs without the need for dissolution and re-nucleation of the aggregates. Formation of thioflavin T-binding and beta-structured aggregates is substantially more rapid than unfolding of the native state, indicating that the initial aggregation process promotes formation of amyloid structures. Taken together, these findings suggest a mechanism of amyloid formation that may have physiological relevance and in which the amyloid structures result from reorganisation of the molecular interactions within the initially formed non-amyloid aggregates. (c) 2005 Elsevier Ltd. All rights reserved.


Volume: 351(4)      Pages from: 910  to: 922

More Information: This work was supported by grants from the European Union (Project number HPRN-CT-2002-00241), the Italian MIUR (FIRB Project number RBAU015B47_001 and PRIN Project number 2003025755_003; L. 449/97, Sector “Genomica funzionale”, project “Strutture ed interazioni molecolari di prodotti genici”), the Wellcome Trust, the Leverhulme Trust, the Cassa di Risparmio di Firenze (Project numbers 2003.437 and 2003.2029) and the Compagnia di San Paolo (Project number 2003.727).
KeyWords: amyloid; amyloid beta protein; congo red; thioflavine, article; beta sheet; circular dichroism; controlled study; enzyme activity; enzyme inactivation; infrared spectroscopy; kinetics; light scattering; nonhuman; priority journal; protein aggregation; protein analysis; protein conformation; protein folding; protein protein interaction; Sulfolobus solfataricus; transmission electron microscopy, Acid Anhydride Hydrolases; Amyloid; Anilino Naphthalenesulfonates; Archaeal Proteins; Circular Dichroism; Congo Red; Kinetics; Microscopy, Electron; Models, Molecular; Multiprotein Complexes; Protein Binding; Protein Structure, Secondary; Recombinant Proteins; Spectroscopy, Fourier Transform Infrared; Sulfolobus solfataricus; Trifluoroethanol; Troponin T, Sulfolobus solfataricus
DOI: 10.1016/j.jmb.2005.06.043

Citations: 111
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