Polymorphism of Alpha-Synuclein Amyloid Fibrils Depends on Ionic Strength and Protein Concentration

Abstract

Protein aggregate formation is linked with multiple amyloidoses, including Alzheimer‘s and Parkinson‘s diseases. Currently, the understanding of such fibrillar structure formation and propagation is still not sufficient, the outcome of which is a lack of potent, anti-amyloid drugs. The environmental conditions used during in vitro protein aggregation assays play an important role in determining both the aggregation kinetic parameters, as well as resulting fibril structure. In the case of alpha-synuclein, ionic strength has been shown as a crucial factor in its amyloid aggregation. In this work, we examine a large sample size of alpha-synuclein aggregation reactions under thirty different ionic strength and protein concentration combinations and determine the resulting fibril structural variations using their dye-binding properties, secondary structure and morphology. We show that both ionic strength and protein concentration determine the structural variability of alpha-synuclein amyloid fibrils and that sometimes even identical conditions can result in up to four distinct types of aggregates.

α‐Synuclein phase separation and amyloid aggregation are modulated by C‐terminal truncations

“…The NaCl concentrations we used in the study are similar to previous reports for the LLPS of a-Syn and other proteins like tau [53][54][55]57,59,60]. These results are consistent with previous findings that a-Syn forms extended conformations in moderate NaCl concentrations, leading to enhancement of fibrillar aggregation [30][31][32].…”

“…The aggregation of a-Syn has been extensively studied in vitro. A variety of environmental conditions can influence the kinetics of a-Syn aggregation [24][25][26][27][28][29][30][31][32]. For example, elevated protein concentration facilitates a-Syn aggregation.…”

“…Mildly acidic pH values can increase the rate of secondary nucleation [29,30]. Furthermore, the ionic strength also plays a significant role in a-Syn aggregation, and the efficient fibrillation requires a moderate NaCl concentration [30][31][32].…”

Amyloid Formation in Nanoliter Droplets

“…This finding implies that amyloids produced in a tight space in the body will exhibit significantly different properties from those produced in a bulk solution [ 31 ]. To date, studies of amyloid fibril polymorphisms have primarily focused on the characteristics of solutions that constitute the surrounding environment, such as the pH, ion strength, additives, and stirring rate, but not the system volume [ 25 , 32 , 33 , 34 , 35 , 36 ]. Considering the results of this study, it was clear that not only the characteristics of the solution, but also the shape and size of the space affect the behavior of amyloid molecules.…”

Interactions between S100A9 and Alpha-Synuclein: Insight from NMR Spectroscopy

“…The fibril structure is related to distinct neurodegenerative diseases [ 18 , 19 ]. The aggregation of -syn is a stochastic process, which depends on the experimental conditions such as solution pH [ 20 ], ionic strength [ 21 , 22 ], protein concentration [ 23 ], or the presence of other amyloid proteins [ 24 , 25 , 26 , 27 , 28 ], including S100A9 [ 29 ] or non-amyloid proteins [ 30 , 31 , 32 ]. Furthermore, 65 proteins in Lewy’s bodies were shown to interact with -syn [ 33 ].…”