Systematic Comparison of the Effects of Alpha-synuclein Mutations on Its Oligomerization and Aggregation (original) (raw)
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Proceedings of the National Academy of Sciences, 2005
Abnormal biology of ␣-synuclein (␣-Syn) is directly implicated in the pathogenesis of Parkinson's disease and other ␣-synucleinopathies. Herein, we demonstrate that C-terminally truncated ␣-Syn (␣-Syn⌬C), enriched in the pathological ␣-Syn aggregates, is normally generated from full-length ␣-Syn independent of ␣-Syn aggregation in brains and in cultured cells. The accumulation of ␣-Syn⌬C is enhanced in neuronal cells as compared with nonneuronal cells. Significantly, the expression of familial Parkinson's disease-linked mutant ␣-Syn is associated with the enhanced cellular accumulation of ␣-Syn⌬C. Moreover, substoichiometric amounts of ␣-Syn⌬C enhance the in vitro aggregation of the more abundant full-length ␣-Syn. Finally, cases of ␣-synucleinopathy exhibit increases in the total soluble ␣-Syn and a higher proportion of soluble ␣-Syn⌬C, a condition favoring the aggregation of ␣-Syn.
Genetic and Pharmacological Modulation of Alpha-Synuclein Aggregation
2017
Several neurodegenerative disorders, such as Parkinson’s disease (PD), are characterized by the deposition of misfolded and aggregated forms of a particular protein in different areas of the brain. Understanding the molecular mechanisms of neurodegenerative diseases are extremely important to prevent and stop such debilitate diseases. PD is a movement related disorder that primarily affects aged individuals, but mutations on alpha-Synuclein (aSyn) gene (SNCA) have been identified in an early- and juvenile-onset of the disease. aSyn is a small an intrinsically disorder protein, that binds to membrane and lipids. It is the major component of Lewy Bodies (LBs) and Lewy Neurites (LN) in the surviving neurons in parkinsonian brains. However, the molecular mechanisms that lead to the selective degeneration of dopaminergic neurons from the substantia nigra pars compacta are still unclear. aSyn aggregation is an important process for the pathology. In pathological conditions, aSyn aggregate...
The effects of the novel A53E alpha- synuclein mutation on its oligomerization and aggregation
α-synuclein (aSyn) is associated with both sporadic and familial forms of Parkinson's disease (PD), the second most common neurodegenerative disorder after Alzheimer's disease. In particular, multiplications and point mutations in the gene encoding for aSyn cause familial forms of PD. Moreover, the accumulation of aSyn in Lewy Bodies and Lewy neurites in disorders such as PD, dementia with Lewy bodies, or multiple system atrophy, suggests aSyn misfolding and aggregation plays an important role in these disorders, collectively known as synucleinopathies. The exact function of aSyn remains unclear, but it is known to be associated with vesicles and membranes, and to have an impact on important cellular functions such as intracellular trafficking and protein degradation systems, leading to cellular pathologies that can be readily studied in cell-based models. Thus, understanding the molecular effects of aSyn point mutations may provide important insight into the molecular mechanisms underlying disease onset. We investigated the effect of the recently identified A53E aSyn mutation. Combining in vitro studies with studies in cell models, we found that this mutation reduces aSyn aggregation and increases proteasome activity, altering normal proteostasis. We observed that, in our experimental paradigms, the A53E mutation affects specific steps of the aggregation process of aSyn and different cellular processes, providing novel ideas about the molecular mechanisms involved in synucleinopathies.
Both Familial Parkinson's Disease Mutations Accelerate alpha -Synuclein Aggregation
Journal of Biological Chemistry, 1999
Parkinson's disease (PD) is a neurodegenerative disorder that is pathologically characterized by the presence of intracytoplasmic Lewy bodies, the major component of which are filaments consisting of ␣-synuclein. Two recently identified point mutations in ␣-synuclein are the only known genetic causes of PD, but their pathogenic mechanism is not understood. Here we show that both wild type and mutant ␣-synuclein form insoluble fibrillar aggregates with antiparallel -sheet structure upon incubation at physiological temperature in vitro. Importantly, aggregate formation is accelerated by both PD-linked mutations. Under the experimental conditions, the lag time for the formation of precipitable aggregates is about 280 h for the wild type protein, 180 h for the A30P mutant, and only 100 h for the A53T mutant protein. These data suggest that the formation of ␣-synuclein aggregates could be a critical step in PD pathogenesis, which is accelerated by the PD-linked mutations.
Proceedings of the National Academy of Sciences, 2005
Abnormal biology of α-synuclein (α-Syn) is directly implicated in the pathogenesis of Parkinson's disease and other α-synucleinopathies. Herein, we demonstrate that C-terminally truncated α-Syn (α-SynΔC), enriched in the pathological α-Syn aggregates, is normally generated from full-length α-Syn independent of α-Syn aggregation in brains and in cultured cells. The accumulation of α-SynΔC is enhanced in neuronal cells as compared with nonneuronal cells. Significantly, the expression of familial Parkinson's disease-linked mutant α-Syn is associated with the enhanced cellular accumulation of α-SynΔC. Moreover, substoichiometric amounts of α-SynΔC enhance the in vitro aggregation of the more abundant full-length α-Syn. Finally, cases of α-synucleinopathy exhibit increases in the total soluble α-Syn and a higher proportion of soluble α-SynΔC, a condition favoring the aggregation of α-Syn. Collectively, our results indicate that the biology behind the generation and accumulation o...
The Parkinson’s Disease-Associated H50Q Mutation Accelerates α-Synuclein Aggregation in Vitro
Biochemistry, 2013
α-Synuclein (α-Syn) aggregation is directly linked with Parkinson's disease (PD) pathogenesis. Here, we analyzed the aggregation of newly discovered α-Syn missense mutant H50Q in vitro and found that this mutation significantly accelerates the aggregation and amyloid formation of α-Syn. This mutation, however, did not alter the overall secondary structure as suggested by two-dimensional nuclear magnetic resonance and circular dichroism spectroscopy. The initial oligomerization study by cross-linking and chromatographic techniques suggested that this mutant oligomerizes to an extent similar to that of the wild-type α-Syn protein. Understanding the aggregation mechanism of this H50Q mutant may help to establish the aggregation and phenotypic relationship of this novel mutant in PD.
Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics, 2019
The misfolding and aggregation of alpha-synuclein (aSyn) are thought to be central events in synucleinopathies. The physiological function of aSyn has been related to vesicle binding and trafficking, but the precise molecular mechanisms leading to aSyn pathogenicity are still obscure. In cell models, aSyn does not readily aggregate, even upon overexpression. Therefore, cellular models that enable the study of aSyn aggregation are essential tools for our understanding of the molecular mechanisms that govern such processes. Here, we investigated the structural features of SynT, an artificial variant of aSyn that has been widely used as a model of aggregation in mammalian cell systems, since it is more prone to aggregation than aSyn. Using Nuclear Magnetic Resonance (NMR) spectroscopy we performed a detailed structural characterization of SynT through a systematic comparison with normal, unmodified aSyn. Interestingly, we found that the conformations adopted by SynT resemble those described for the unmodified protein, demonstrating the usefulness of SynT as a model for aSyn aggregation. However, subtle differences were observed at the N-terminal region involving transient intra and/or intermolecular interactions that are known to regulate aSyn aggregation. Importantly, our results indicate that disturbances in the N-terminal region of SynT, and the consequent decrease in membrane binding of the modified protein, might contribute to the observed aggregation behavior of aSyn, and validate the use of SynT, one of the few models of aSyn aggregation in cultured cells.
Journal of Biological Chemistry, 1999
Parkinson's disease (PD) is a neurodegenerative disorder that is pathologically characterized by the presence of intracytoplasmic Lewy bodies, the major components of which are filaments consisting of ␣-synuclein. Two recently identified point mutations in ␣-synuclein are the only known genetic causes of PD. ␣-Synuclein fibrils similar to the Lewy body filaments can be formed in vitro, and we have shown recently that both PDlinked mutations accelerate their formation. This study addresses the mechanism of ␣-synuclein aggregation: we show that (i) it is a nucleation-dependent process that can be seeded by aggregated ␣-synuclein functioning as nuclei, (ii) this fibril growth follows first-order kinetics with respect to ␣-synuclein concentration, and (iii) mutant ␣-synuclein can seed the aggregation of wild type ␣-synuclein, which leads us to predict that the Lewy bodies of familial PD patients with ␣-synuclein mutations will contain both, the mutant and the wild type protein. Finally (iv), we show that wild type and mutant forms of ␣-synuclein do not differ in their critical concentrations. These results suggest that differences in aggregation kinetics of ␣-synucleins cannot be explained by differences in solubility but are due to different nucleation rates. Consequently, ␣-synuclein nucleation may be the rate-limiting step for the formation of Lewy body ␣-synuclein fibrils in Parkinson's disease.