Scutellarin inhibits the uninduced and metal-induced aggregation of α-Synuclein and disaggregates preformed fibrils: implications for Parkinson's disease

Abstract

The aggregation of the protein alpha synuclein (α-Syn), a known contributor in Parkinson's disease (PD) pathogenesis is triggered by transition metal ions through occupational exposure and disrupted metal ion homeostasis. Naturally occurring small molecules such as polyphenols have emerged as promising inhibitors of α-Syn fibrillation and toxicity and could be potential therapeutic agents against PD. Here, using an array of biophysical tools combined with cellular assays, we demonstrate that the novel polyphenolic compound scutellarin efficiently inhibits the uninduced and metal-induced fibrillation of α-Syn by acting at the nucleation stage and stabilizes a partially folded intermediate of α-Syn to form SDS-resistant, higher-order oligomers (∼680 kDa) and also disaggregates preformed fibrils of α-Syn into similar type of higher-order oligomers. ANS binding assay, fluorescence lifetime measurements and cell-toxicity experiments reveal scutellarin-generated oligomers as compact, low hydrophobicity structures with modulated surface properties and significantly reduced cytotoxicity than the fibrillation intermediates of α-Syn control. Fluorescence spectroscopy and isothermal titration calorimetry establish the binding between scutellarin and α-Syn to be non-covalent in nature and of moderate affinity (Ka ∼ 105 M−1). Molecular docking approaches suggest binding of scutellarin to the residues present in the NAC region and C-terminus of monomeric α-Syn and the C-terminal residues of fibrillar α-Syn, demonstrating inhibition of fibrillation upon binding to these residues and possible stabilization of the autoinhibitory conformation of α-Syn. These findings reveal interesting insights into the mechanism of scutellarin action and establish it as an efficient modulator of uninduced as well as metal-induced α-Syn fibrillation and toxicity.

Crocin Inhibits the Fibrillation of Human α-synuclein and Disassembles Mature Fibrils: Experimental Findings and Mechanistic Insights from Molecular Dynamics Simulation

“…On the other hand, the strong affinity of crocin to the NAC region could be hampering the interactions and rearrangements required for the β-sheet aggregation, thus suppressing the hαS fibrillogenesis. Furthermore, as intramolecular contacts between the C-terminal and NAC regions are considered to protect the amyloidogenic NAC domain from intermolecular interactions that lead to aggregation under native conditions, binding of crocin to both these domains could reinforce or prolong autoinhibitory contacts, contributing to aggregation inhibition, as suggested by some researchers. , …”

“…Furthermore, as intramolecular contacts between the C-terminal and NAC regions are considered to protect the amyloidogenic NAC domain from intermolecular interactions that lead to aggregation under native conditions, 58 binding of crocin to both these domains could reinforce or prolong autoinhibitory contacts, contributing to aggregation inhibition, as suggested by some researchers. 18,72 Crocin Molecules Resided Longitudinally along the Fibril Axis onto the Edges of the Inter-protofilament Interface of the hαS Fibril. A recently solved cryo-electron microscopybased high-resolution structure of the hαS fiber (PDB id: 6A6B) was utilized as the structural template to examine the hαS fibril−crocin interaction.…”

Effects of α-Synuclein-Associated Post-Translational Modifications in Parkinson’s Disease

“…α-Syn controls the aggregation and release of synaptic vesicles synapses and stabilizes the electron transport chain protein on the mitochondrial membrane together with cardiolipin. Furthermore, factors such as oxidative stress, , proteolysis, , fatty acid concentration, , phospholipids and metal ions − regulate the structure of α-syn, leading to different forms of proteins, including oligomers and fibers, that can develop into cytoplasmic inclusions.…”

Natural and Synthetic Derivatives of Hydroxycinnamic Acid Modulating the Pathological Transformation of Amyloidogenic Proteins

“…The utilization of ensemble docking looks more promising when potential ligands are docked not to the one “initial” structure, but to the ensemble of conformations gathered by molecular dynamic simulations [ 132 ]. This approach was used, for example, to predict the binding site of phenolic compounds (noradrenaline and scutellarin) [ 133 , 134 ]. Difficulties of this approach include the complexity of the method and the fact that the quality of the result depends on how close to reality the ensemble of conformations is.…”