Primary Nucleation of Polymorphic α-Synuclein Dimers Depends on Copper Concentrations and Definite Copper-Binding Site

Abstract

The primary nucleation process of α-synuclein (AS) that forms toxic oligomeric species is the early stage of the pathological cause of Parkinson's disease. It is well-known that copper influences this primary nucleation process. While significant efforts have been made to solve the structures of polymorphic AS fibrils, the structures of AS oligomers and the copper-bound AS oligomers at the molecular level and the effect of copper concentrations on the primary nucleation are elusive. Here, we propose and demonstrate new molecular mechanism pathways of primary nucleation of AS that are tuned by distinct copper concentrations and by a specific copper-binding site. We present the polymorphic AS dimers bound to different copper-binding sites at the atomic resolution in high- and low-copper concentrations, using extensive molecular dynamics simulations. Our results show the complexity of the primary nucleation pathways that rely on the copper concentrations and the copper binding site. From a broader perspective, our study proposes a new strategy to control the primary nucleation of other toxic amyloid oligomers in other neurodegenerative diseases.

Keywords: Parkinson’s disease; amyloids; metal ions; neurodegenerative diseases; oligomers; polymorphism; protein aggregation; self-assembly; α-synuclein.

Figure 1

Final simulated Cu²⁺-AS_1–140 dimer models at low copper concentrations: A1–A3, and B1–B3; and at high copper concentrations: A4–A6, and B4–B6. Descriptions of the models are detailed in Table 1.

Figure 2

Final simulated Cu²⁺-AS_1–140 dimer models at low copper concentrations: C1–C3, and D1–D3; and at high copper concentrations: C4–C6, and D4–D6. Descriptions of the models are detailed in Table 1.

Figure 3

The percentages by which Cu²⁺ ions bind to each residue along the MD simulations, for the binding sites. (a) Met1, Asp2, and Met5; (b) His50; and (c) Asp119, Asp121, Asn122, and Glu123.

Figure 4

The percentages of the secondary structure along the sequence of AS in Cu²⁺-free polymorphic AS_1–140 dimers (models A, B, C, and D) [5] and in Cu²⁺-bound polymorphic AS_1–140 dimers (models A1–A6, B1–B6, C1–C6, and D1–D6). The sequence of the NAC domain is indicated in green rectangles.

Figure 5

(a) Conformational energies (Top) and populations (Bottom) of the Cu²⁺-bound AS_1–140 dimer models at low copper concentrations, and (b) conformational energies (Top) and populations (Bottom) of the Cu²⁺-bound AS_1–140 dimer models at high copper concentrations.

Figure 6

Hydrophobic interaction occurrence NAC contact maps between two Cu²⁺-bound AS monomers within the dimer models at low copper concentrations (models A1–A3, B1–B3, C1–C3, D1–D3).

Figure 7

Hydrophobic interaction occurrence NAC contact maps between two Cu²⁺-bound AS monomers within the dimer models at high copper concentrations (models A4–A6, B4–B6, C4–C6, D4–D6).

Figure 8

Root-mean-square fluctuation (RMSF) values of along the sequence of AS in Cu²⁺-free polymorphic AS_1–140 dimers (models A, B, C, and D) [5] and in Cu²⁺-bound polymorphic AS_1–140 dimers (models A1–A6, B1–B6, C1–C6, and D1–D6).

Figure 9

Proposed mechanisms for Cu²⁺-binding sites in polymorphic AS dimers models A (orange), model B (pink), model C (blue), and model D (purple): The Cu²⁺ concentrations affect the specific Cu²⁺-binding site: (a) At low Cu²⁺ concentration, the binding is preferred by Asp2, whereas, at high Cu²⁺ concentration, additional residues participate in the Cu²⁺-binding. (b) Both at low and high Cu²⁺ concentrations, the His50 binds to Cu²⁺ and to further residues from both N- and C-termini domains. (c) Both at low and high Cu²⁺ concentrations, a common Cu²⁺-binding site appears at the C-termini: Asp119, Asp121, and Glu123 (excluding in model C).