Loading...
Research Project
Untitled
Funder
Authors
Publications
The V30M amyloidogenic mutation decreases the rate of refolding kinetics of the tetrameric protein transthyretin
Publication . Jesus, Catarina S. H.; Vaz, Daniela C.; Saraiva, Maria J. M.; Brito, Rui M. M.
Transthyretin (TTR) is a homotetrameric protein implicated in several amyloid diseases. The mechanism by which TTR is converted into elongated fibrillar assemblies has been extensively investigated, and numerous studies showed that dissociation of the native tetrameric structure into partially unfolded monomeric species precedes amyloid formation. The small differences observed in the crystal structures of different TTR variants, as well as the thermodynamics and kinetics of tetramer dissociation, do not seem to completely justify the amyloidogenic potential of different TTR variants. With this in mind, we have studied the refolding kinetics of WT-TTR and its most common amyloidogenic variant V30M-TTR, monitoring changes in intrinsic tryptophan fluorescence at different urea and protein concentrations. Our results demonstrate that the in vitro refolding
mechanisms of WT- and V30M-TTR are similar, involving a dimeric intermediate. However, there are large differences in the refolding rate constants for the two variants, specially close to physiological conditions. Interestingly, tetramer formation occurs at a much slower rate in the amyloidogenic variant V30M-TTR than in WT-TTR, which in the in vivo setting may promote the accumulation of monomeric species in the extracellular environment, resulting in higher susceptibility for aggregation and amyloid formation instead of spontaneous refolding.
A New Folding Kinetic Mechanism for Human Transthyretin and the Influence of the Amyloidogenic V30M Mutation
Publication . Jesus, Catarina S. H.; Almeida, Zaida L.; Vaz, Daniela C.; Faria, Tiago Q.; Brito, Rui M. M.
Protein aggregation into insoluble amyloid fibrils is the hallmark of several
neurodegenerative diseases, chief among them Alzheimer’s and Parkinson’s. Although caused
by different proteins, these pathologies share some basic molecular mechanisms with familial
amyloidotic polyneuropathy (FAP), a rare hereditary neuropathy caused by amyloid formation
and deposition by transthyretin (TTR) in the peripheral and autonomic nervous systems. Among the
amyloidogenic TTR mutations known, V30M-TTR is the most common in FAP. TTR amyloidogenesis
(ATTR) is triggered by tetramer dissociation, followed by partial unfolding and aggregation of the
low conformational stability monomers formed. Thus, tetramer dissociation kinetics, monomer
conformational stability and competition between refolding and aggregation pathways do play
a critical role in ATTR. Here, we propose a new model to analyze the refolding kinetics of WT-TTR
and V30M-TTR, showing that at pH and protein concentrations close to physiological, a two-step
mechanism with a unimolecular first step followed by a second-order second step adjusts well to the
experimental data. Interestingly, although sharing the same kinetic mechanism, V30M-TTR refolds at
a much slower rate than WT-TTR, a feature that may favor the formation of transient species leading
to kinetic partition into amyloidogenic pathways and, thus, significantly increasing the probability of
amyloid formation in vivo.
Organizational Units
Description
Keywords
Contributors
Funders
Funding agency
Fundação para a Ciência e a Tecnologia
Funding programme
SFRH
Funding Award Number
SFRH/BD/43896/2008