A Combined Approach to Extract Rotational Dynamics of Globular Proteins Undergoing Liquid–Liquid Phase Separation

The Journal of Physical Chemistry B, Journal Year: 2025, Volume and Issue: unknown

Published: Jan. 16, 2025

The formation of protein condensates (droplets) via liquid-liquid phase separation (LLPS) is a commonly observed phenomenon in vitro. Changing the environmental properties with cosolutes, molecular crowders, partners, temperature, pressure, etc. has been shown to favor or disfavor droplets by fine-tuning water-water, water-protein, and protein-protein interactions. Therefore, these their spatiotemporal are likely be important also cellular context at existing expression levels. One key physicochemical biomolecules impacted crowding diffusion, which determines viscoelastic behavior condensates. Here, we investigate change rotational diffusion γD-crystallin, undergoing LLPS vitro aqueous solutions absence presence cosolutes. We studied its dynamics using simulations (MD), electron paramagnetic resonance (EPR) spectroscopy, fluorescence spectroscopy. MD performed under dilute crowded conditions show that crystallin water retarded 1 2 orders magnitude condensed phase. To obtain phase, used anisotropy extract retardation factor spin-labeled γD-crystallin proteins as EPR viscosity nanoprobes. Aided nanoruler calibrated increasing sucrose concentrations, validated predicted simulations. This study underlines predictive power showcases use sensitive nanoprobe biomolecular

Language: Английский

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Protein Crowding Effects on Hydration Water Dynamics

The Journal of Physical Chemistry Letters, Journal Year: 2025, Volume and Issue: 16(9), P. 2340 - 2347

Published: Feb. 24, 2025

We propose a time-resolved optical Kerr effect study of the structural and vibrational dynamics hydration water surrounding lysozyme on very fast time scale. Measurements as function concentration make it possible to distinguish contribution from that both bulk protein. Our results provide experimental evidence existence two water, associated with hydrogen bond exchange relaxation process reorganization molecules induced by protein fluctuations. Likewise, we evaluated layer at subpicosecond scales. measurements properties reveal presence crossover point specific concentration. This marks transition between clustering regimes distinct characteristics establishes threshold for crowding.

Language: Английский

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Tuning biological processes via co-solutes: from single proteins to protein condensates – the case of α-elastin condensation

Chemical Science, Journal Year: 2025, Volume and Issue: unknown

Published: Jan. 1, 2025

Protein condensates as membrane-less compartments play a pivotal role in cellular processes. The stabilization of protein condensation can be tuned using cosolutes which directly impact biological function. In this study, we report the result rigorous study influence changes on hydration entropy and enthalpy upon condensate formation, by means THz-calorimetry. Our results unveil quantitative insights into fine tuning free energy imbalance, via hydrophobic/entropic hydrophilic/enthalpic cosolute-mediated or destabilization condensates. These shed new light regulatory potential co-solutes within cells, to tune Liquid-Liquid Phase Separation (LLPS). Furthermore, demonstrate transferability underlying molecular concepts cosolute addition two fundamental processes: folding denaturation. This provides blueprint for controlled modulating LLPS additions, with promising implications both medical applications.

Language: Английский

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Deciphering driving forces of biomolecular phase separation from simulations

Current Opinion in Structural Biology, Journal Year: 2025, Volume and Issue: 92, P. 103026 - 103026

Published: March 8, 2025

The formation and modulation of biomolecular condensates as well their structural dynamic properties are determined by an intricate interplay different driving forces, which down at the microscopic scale involve molecular interactions biological macromolecules surrounding solvent ions. Molecular simulations increasingly used to provide detailed insights into various processes thermodynamic forces play, thereby yielding mechanistic understanding aiding interpretation experiments level individual amino acid residues or even atoms. Here we summarize recent advances in field biocondensate with a focus on coarse-grained all-atom dynamics (MD) simulations. We highlight possible future challenges concerning computationally efficient physically accurate large complex systems.

Language: Английский

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Backbone-mediated weakening of pairwise interactions enables percolation in peptide-based mimics of protein condensates

Communications Chemistry, Journal Year: 2025, Volume and Issue: 8(1)

Published: April 6, 2025

Biomolecular condensates formed by intrinsically disordered proteins (IDPs) are semidilute solutions. These can be approximated as solutions of blob-sized segments, which peptide-sized motifs. We leveraged the blob picture and molecular dynamics simulations to quantify differences between inter-residue interactions in model compound peptide-based mimics dense versus dilute phases. The all-atom use a polarizable forcefield. In solutions, aromatic residues stronger than cationic residues. This holds Cooperativity within phases enhances pairwise leading finite-sized nanoscale clusters. results for paint different picture. Backbone amides add valence associating molecules. While this phases, it weakens pair doing so concentration-dependent manner. Weakening enables fluidization characterized short-range order long-range disorder. higher afforded peptide backbone generates system-spanning networks. As result, peptides best described percolated network fluids. Overall, our show how backbones enhance while weakening these enable percolation

Language: Английский

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Thioflavin T─a Reporter of Microviscosity in Protein Aggregation Process: The Study Case of α-Synuclein

The Journal of Physical Chemistry Letters, Journal Year: 2024, Volume and Issue: 15(25), P. 6685 - 6690

Published: June 20, 2024

Thioflavin T (ThT) informed microviscosity changes can be used to monitor protein aggregation. Steady-state, time-resolved and lasing spectroscopy were detect transient states in α-synuclein - a associated with Parkinson's disease. The major focus was on the nucleation phase, where conventional ThT fluorescence assay lacks appropriate sensitivity early stage oligomers. Instead, threshold parameters, particular, sensitive detecting Through spectroscopy, change correlating stages of aggregation observed at two wavelengths 405 440 nm. are free dye molecules β-sheet bound molecules. This provides perspective elucidating formed aggregation, critical aspect understanding pathogenesis neurodegenerative diseases. insights from presented study shows potential using as tool studying dynamics.

Language: Английский

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Unraveling the Hydration Dynamics of the ACC1-13K24-ATP System: From Liquid-to-Droplet to-Amyloid Fibril

Biophysical Journal, Journal Year: 2024, Volume and Issue: 123(22), P. 3863 - 3870

Published: Sept. 11, 2024

Language: Английский

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Water-protein Interactions as a Driver of Phase Separation, Biology, and Disease

Biophysical Journal, Journal Year: 2024, Volume and Issue: 123(22), P. 3859 - 3862

Published: Oct. 16, 2024

Language: Английский

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Heterogeneous Slowdown of Dynamics in the Condensate of an Intrinsically Disordered Protein

The Journal of Physical Chemistry Letters, Journal Year: 2024, Volume and Issue: 15(45), P. 11244 - 11251

Published: Nov. 1, 2024

The high concentration of proteins and other biological macromolecules inside biomolecular condensates leads to dense confined environments, which can affect the dynamic ensembles time scales conformational transitions. Here, we use atomistic molecular dynamics (MD) simulations intrinsically disordered low complexity domain (LCD) human fused in sarcoma (FUS) RNA-binding protein study how self-crowding a condensate affects motions protein. We found heterogeneous retardation with respect dilute phase, large-amplitude being strongly slowed by up 2 orders magnitude, whereas small-scale motions, such as local backbone fluctuations side-chain rotations, are less affected. results support notion liquid-like character show that different respond differently environment.

Language: Английский

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Mechanistic Insights into the c-MYC G-Quadruplex and Berberine Binding inside an Aqueous Two-Phase System Mimicking Biomolecular Condensates

The Journal of Physical Chemistry Letters, Journal Year: 2024, Volume and Issue: 15(34), P. 8706 - 8714

Published: Aug. 19, 2024

We investigated the binding between c-MYC G-quadruplex (GQ) and berberine chloride (BCl) in an aqueous two-phase system (ATPS) with 12.3 wt % polyethylene glycol 5.6 dextran, mimicking highly crowded intracellular biomolecular condensates formed via liquid–liquid phase separation. found that ATPS, complex formation is significantly altered, leading to increase affinity a change stoichiometry of respect neat buffer conditions. Thermodynamic studies reveal becomes more thermodynamically favorable ATPS due entropic effects, as strong excluded volume effect inside droplets reduces penalty associated binding. Finally, BCl for GQ higher than those other DNA structures, indicating potential specific interactions. Overall, these findings will be helpful design drugs targeting structures cancer-related biocondensates.

Language: Английский

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