Abstract
In amyloidosis, normally innocuous soluble proteins polymerize to form insoluble fibrils. Amyloid fibril formation and deposition have been associated with a wide range of diseases, including spongiform encephalopathies, Alzheimer's disease, and familial amyloid polyneuropathies (FAP). In certain forms of FAP, the amyloid fibrils are mostly constituted by variants of transthyretin (TTR), a homotetrameric plasma protein implicated in the transport of thyroxine and retinol. The most common amyloidogenic TTR variant is V30M-TTR, and L55P-TTR is the variant associated with the most aggressive form of FAP. Recently, we reported that TTR dissociates to a monomeric species at pH 7.0 and nearly physiological ionic strengths (Quintas, A., Saraiva, M. J., and Brito, R. M. (1997) FEBS Lett. 418, 297-300). Here, we show that the tetramer dissociation is apparently irreversible; and based on intrinsic tryptophan fluorescence and fluorescence quenching experiments, we show that the monomeric species formed upon tetramer dissociation is non-native. We also show, based on 1-anilino-8-naph-thalenesulfonate binding studies, that this monomeric species appears not to behave like a molten globule. These data allowed us to propose a model for TTR amyloidogenesis based on tetramer dissociation occurring naturally under commonly observed physiological solution conditions.
Highlights
In amyloidosis, normally innocuous soluble proteins polymerize to form insoluble fibrils
In certain forms of familial amyloid polyneuropathies (FAP), the amyloid fibrils are mostly constituted by variants of transthyretin (TTR), a homotetrameric plasma protein implicated in the transport of thyroxine and retinol
The solvent-accessible surface area of the Transthyretin Dissociation—Tetrameric and monomeric species of WT- and V30M-TTR were isolated by gel filtration chromatography based on the dissociation behavior of the tetramers, previously reported [16]
Summary
(Received for publication, February 16, 1999, and in revised form, August 25, 1999). From the ‡Centro de Neurociencias de Coimbra, Universidade de Coimbra, 3004-517 Coimbra, Portugal, the §Instituto Superior de Ciencias da Saude Sul, Quinta da Granja, 2825 Monte da Caparica, Portugal, the ʈInstituto de Ciencias Biomedicas de Abel Salazar and the Amyloid Unit, Institute for Molecular and Cellular Biology, Universidade do Porto, 4050 Porto, Portugal, and the **Departamento de Quımica, Faculdade de Ciencias e Tecnologia, Universidade de Coimbra, 3049 Coimbra, Portugal. In certain forms of FAP, the amyloid fibrils are mostly constituted by variants of transthyretin (TTR), a homotetrameric plasma protein implicated in the transport of thyroxine and retinol. We show, based on 1-anilino-8-naph-thalenesulfonate binding studies, that this monomeric species appears not to behave like a molten globule These data allowed us to propose a model for TTR amyloidogenesis based on tetramer dissociation occurring naturally under commonly observed physiological solution conditions. This observation has led to the proposal of a low pH environment as a prerequisite for amyloid formation in vivo [9] These authors proposed that the low pH medium (present for example in the lysosomes) would induce tetramer rearrangement and dissociation to a monomeric amyloidogenic intermediate with altered tertiary structure, which in turn would self-assemble to form amyloid fibrils [10]. Monly observed physiological conditions and consistent with extracellular amyloid deposit formation
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