Crucial role of nonspecific interactions in amyloid nucleation

Abstract

Protein oligomers have been implicated as toxic agents in a wide range of amyloid-related diseases. However, it has remained unsolved whether the oligomers are a necessary step in the formation of amyloid fibrils or just a dangerous byproduct. Analogously, it has not been resolved if the amyloid nucleation process is a classical onestep nucleation process or a two-step process involving prenucleation clusters. We use coarse-grained computer simulations to study the effect of nonspecific attractions between peptides on the primary nucleation process underlying amyloid fibrillization. We find that, for peptides that do not attract, the classical one-step nucleation mechanism is possible but only at nonphysiologically high peptide concentrations. At low peptide concentrations, which mimic the physiologically relevant regime, attractive interpeptide interactions are essential for fibril formation. Nucleation then inevitably takes place through a two-step mechanism involving prefibrillar oligomers. We show that oligomers not only help peptides meet each other but also, create an environment that facilitates the conversion of monomers into the β-sheet-rich form characteristic of fibrils. Nucleation typically does not proceed through the most prevalent oligomers but through an oligomer size that is only observed in rare fluctuations, which is why such aggregates might be hard to capture experimentally. Finally, we find that the nucleation of amyloid fibrils cannot be described by classical nucleation theory: in the two-step mechanism, the critical nucleus size increases with increases in both concentration and interpeptide interactions, which is in direct contrast with predictions from classical nucleation theory.amyloid | protein aggregation | protein oligomers | neurodegeneration | coarse-grained simulations D uring the process of amyloid formation, normally soluble proteins assemble into fibrils that are enriched in β-sheet content and have diameters of a few nanometers and lengths up to several micrometers. This phenomenon has been implicated in a variety of pathogenic processes, including Alzheimer's and Parkinson's diseases, type 2 diabetes, and systemic amyloidoses (1-3). The association with human diseases has largely motivated a long-standing effort to probe the assembly process, and numerous studies have aimed at elucidating the mechanism of amyloid aggregation (4). The basic nature of the aggregation reaction has emerged as a nucleation and growth process (5, 6), where the aggregates are created through a not well-understood primary nucleation event and can grow by recruiting additional peptides or proteins to their ends (7,8). In this paper, we focus on the nature of this primary step in amyloid nucleation and the fundamental initial events that underlie amyloid formation.Amyloidogenic peptides and proteins, when in their nonpathological cellular form, can range in the structures from mainly α-helical to β-sheet and even random coil, whereas the amyloid forms of proteins possess a generic cross-β-structure ...

Mechanisms and rates of nucleation of amyloid fibrils

“…(There is an additional feature: the re-increase of N * for a low ∆ αβ = −18 k B T at very high concentrations, which we do not fully understand and will not be discussing further). These values of 'critical nucleus' size are within the range obtained by the coarse-grained molecular simulations of Sarić et al 29 : from 2 to 14; they also fall within the experimentally observed values (2 to roughly 25) 80 . Notably, the fact that N * is not a constant value in Fig.…”

“…It is suggested that these amorphous micelles serve as intermediates, or initiation states for amyloid nucleation 63 . This generic hypothesis of the role of micelles is indirectly hinted by the increase in β-sheet structural content while losing their original α-helix content 62 , and is further supported by molecular simulation of coarse-grained peptides 29,64 . These facts imply an alternative aggregation pathway for amyloid fibrils: the two-step nucleation mechanism as described in Fig.…”

“…1. However, we find it is sufficient to use a two-state simplification to capture the essence of amyloid aggregation mechanism, as has been suggested in molecular simulations 28,29 : we denote the soluble monomer as 'α-mer' (for its assumed increasing content of α helix when forming micelles), while the β-mer is the monomeric unit of mature amyloid fibrils. We later use this two-state simplification to schematically show two different nucleation mechanisms as an aid to point out the weakness of previous theoretical kinetic models on determining the critical nucleus size.…”

Kinetics of spontaneous filament nucleation via oligomers: Insights from theory and simulation

“…We employ a minimal Monte Carlo computational model that reproduces fibril nucleation, as described in our previous work 45,104 . Briefly, the model accounts for the fact that amyloidogenic peptides and proteins exist in minimally two states: a state in solution (denoted "s") that can form disordered oligomers, and a higher free-energy state that can form the β-sheet enriched fibrils (denoted "β") 104,109 .…”

“…The "s" -"β" interaction was set to sβ = ss + 1kT , as in our previous work 104 . Throughout the text k denotes Boltzmann's constant and T the temperature.…”