Negatively Charged α-Synuclein Condensate Modulates Partitioning of Molecules

α-Synuclein (αSyn) aggregation via liquid-liquid phase separation has recently emerged as a crucial mechanism underlying amyloid fibril formation implicated in Parkinson’s disease. However, comprehensive investigations of the physico-chemical properties of αSyn condensate remains incomplete. Here, we showed that αSyn condensate possesses a highly negative electrostatic potential, which is manifested as differential partitioning of dye-labeled αSyn as well as fluorescent molecules by an order of magnitude depending on their net charges. Consistent with this, the phase separation potency of αSyn is governed by a delicate balance between self-association of αSyn and electrostatic repulsion, hence is antagonized by excess negative charge. We further demonstrated that, in differentiated neuron-like SH-SY5Y cells, αSyn also forms negatively charged condensate. Our results highlighted the significant impact of charged fluorescent tag on the partitioning property as well as the phase-separated states of αSyn condensate, hence calling for close examination of the electrostatic property of other biomolecular condensates.