Analysis of α-Synuclein LLPS in Neurodegenerative Diseases

α-Synuclein is a naturally unstructured protein that performs many physiological functions in neurons, including vesicular transport. Studies have shown that α-Synuclein displays protein cohesion with liquid-like behavior in both cellular and animal models, performing liquid-liquid phase separation (LLPS) through simple condensates of predominantly hydrophobic interactions. α-Synuclein amyloid aggregates have been observed within neurons in Parkinson's disease (PD) and other synucleinopathies, but the heterogeneous and transient nature of the oligomers formed in the early stages of the aggregation pathway has limited their detailed study in understanding structure-toxicity relationships. In addition, the exact mechanism by which α-Synuclein amyloid aggregation occurs intracellularly remains unclear.

Fig. 1. In vitro LLPS of α-Synuclein. (Mukherjee S, et al., 2023)

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LLPS of α-Synuclein occurs during the nucleation step of its aggregation, undergoing irreversible liquid- to solid-phase transformation into different pathological aggregates, including large protofibrils, and various pre-protofibrillar oligomeric species. These aggregated α-synaptic nucleoprotein species are hallmarks of PD neuropathology. Synaptic nucleopathy can be avoided by preventing the accumulation of α-Synuclein.

Here, CD BioSciences offers professional services to analyze LLPS of α-Synuclein in PD. Our technical team can capture the current state of the science and comprehensively analyze the exact molecular structure and function of α-Synuclein in the progression of neurodegenerative diseases.

We offer the following physicochemical strategies to characterize the LLPS of α-Synuclein.

• We provide turbidimetric quantification of quantified α-Synuclein to confirm the electrostatic nature of the major interactions associated with its stability and to assess the effect of different polymer ratios on the LLPS process.
• We have sophisticated high-throughput techniques to simultaneously monitor α-Synuclein concentration and ThT binding signal in the dilute phase during α-Synuclein liquid-solid phase transition. We aim to explore all intermediate species in the LLPS-mediated α-Synuclein aggregation pathway and their potential cytotoxicity.
• Our laboratory has advanced microscopy and spectroscopy platforms to characterize the viscoelastic transition of α-Synuclein droplets over time at individual droplet resolution.

Our experts develop multiple means to modulate α-Synuclein LLPS to prevent aggregation, including:

• Altering experimental conditions (such as concentration, pH).
• Modulating the LLPS behavior of the protein through family mutations and primary sequence modifications of α-Synuclein by PTMs.
• Analyzing the effects of other PD-related covalent modifications (e.g. oxidative modifications, nitration, advanced glycosylation, etc.) on α-Synuclein LLPS.
• Regulation of α-Synuclein LLPS by metal ions such as Mn²⁺ and Ca²⁺.
• Reducing the neuronal level of α-Synuclein by silencing of SNCA gene.
• Increasing aggregated α-Synuclein clearance by stimulating autophagy or proteasome activity.

Furthermore, we systematically integrate these regulatory approaches with proteomic, transcriptomic, imaging, genetic, and epidemiological data to explore the pathogenesis of abnormal phase changes in α-Synuclein and to develop potential therapeutic targets to prevent and treat these neurodegenerative diseases.

CD BioSciences aims to analyze the regulation of α-Synuclein LLPS to develop potential therapeutic approaches for neurodegenerative diseases. If you are interested in our services, please feel free to contact us.