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We have investigated the structural evolution in solutions of the intrinsically disordered protein, α-synuclein, as a function of protein concentration and added salt concentration. Accounting for electrostatic and excluded volume interactions based on the protein sequence, our Langevin dynamics simulations reveal that α-synuclein molecules assemble into aggregates and percolated structures with a spontaneous selection of a dominant structure characteristic of microphase separation. This microphase assembly is mainly driven by electrostatic interactions between the residues in N-terminal and C-terminal of the protein molecules, and presence of salt loosens the compactness of the microstructures. We have quantified the features of the spontaneously formed microstructures using interchain radial distribution functions, and experimentally measurable inter-residue contact maps and static structure factors. Our results are in contrast to the commonly hypothesized mechanism of liquid–liquid phase separation (LLPS) for the formation of droplets in solutions of intrinsically disordered proteins, opening a new paradigm to understand the birth and structure of membraneless organelles. In general, construction of phase diagrams of intrinsically disordered proteins and other biomacromolecular systems needs to incorporate features of microphase separation into other mechanisms of macrophase separation and percolation.
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This content will become publicly available on December 1, 2025
Solutes unmask differences in clustering versus phase separation of FET proteins
Phase separation and percolation contribute to phase transitions of multivalent macromolecules. Contributions of percolation are evident through the viscoelasticity of condensates and through the formation of heterogeneous distributions of nano- and mesoscale pre-percolation clusters in sub-saturated solutions. Here, we show that clusters formed in sub-saturated solutions of FET (FUS-EWSR1-TAF15) proteins are affected differently by glutamate versus chloride. These differences on the nanoscale, gleaned using a suite of methods deployed across a wide range of protein concentrations, are prevalent and can be unmasked even though the driving forces for phase separation remain unchanged in glutamate versus chloride. Strikingly, differences in anion-mediated interactions that drive clustering saturate on the micron-scale. Beyond this length scale the system separates into coexisting phases. Overall, we find that sequence-encoded interactions, mediated by solution components, make synergistic and distinct contributions to the formation of pre-percolation clusters in sub-saturated solutions, and to the driving forces for phase separation.
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- Award ID(s):
- 2227268
- PAR ID:
- 10564799
- Publisher / Repository:
- Nature Publishing Group
- Date Published:
- Journal Name:
- Nature Communications
- Volume:
- 15
- Issue:
- 1
- ISSN:
- 2041-1723
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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