Department of Engineering Mechanics, Hohai University, Nanjing 210098, China.
Department of Physics and Astronomy, Clemson University, Clemson, South Carolina 29634, United States.
J Chem Inf Model. 2024 Oct 14;64(19):7590-7601. doi: 10.1021/acs.jcim.4c00943. Epub 2024 Sep 29.
Liquid-liquid phase separation (LLPS) of transactive response DNA-binding protein of 43 kDa (TDP-43), which exerts multiple functions in the splicing, trafficking, and stabilization of RNA, mediates the formation of membraneless condensates with crucial physiological roles, while its aberrant LLPS is linked to multiple neurodegenerative diseases. However, due to the heterogeneous and dynamic nature of LLPS, major gaps remain in understanding the precise intermolecular interactions driving LLPS and how specific mutations alter LLPS dynamics. Here, we investigated the molecular mechanisms underlying the LLPS of the TDP-43 low-complexity domain (LCD) by simulating the dimerization process using all-atom discrete molecular dynamics with microsecond-long simulations. Our results showed that the TDP-43 LCD was intrinsically disordered, with helical structures consistent with prior nuclear magnetic resonance studies. Phase separation propensity was assessed by simulating the dimerization of the TDP-43 LCD and four mutants, showing that A321G, W334G, and M337V inhibited self-association, while G335D promoted it, fully consistent with experimental reports. During the dimerization process, two peptides experienced both elastic and nonelastic collisions, and the self-associated dimer featured both high- and low-contact states. These results suggested that the dimerization process of the TDP-43 LCD was accordingly dynamic and heterogeneous. Additionally, we identified crucial regions containing hydrophobic clusters and aromatic residues in the N-terminus, central region, and C-terminus that were essential for the self-association of the TDP-43 LCD. These residues with high binding affinities can act as stickers to form peptide networks in LLPS. Together, our simulation provides a comprehensive picture of the intermolecular interactions driving the phase separation of the TDP-43 LCD, offering insights into both physiological functions and pathological mechanisms.
液-液相分离 (LLPS) 的转激活反应 DNA 结合蛋白 43 kDa (TDP-43)，它在剪接、运输和 RNA 稳定中发挥多种功能，介导具有关键生理作用的无膜液滴的形成，而其异常的 LLPS 与多种神经退行性疾病有关。然而，由于 LLPS 的异质性和动态性质，对于驱动 LLPS 的精确分子间相互作用以及特定突变如何改变 LLPS 动力学的理解仍存在很大差距。在这里，我们通过使用微秒长模拟的全原子离散分子动力学模拟二聚化过程来研究 TDP-43 低复杂度结构域 (LCD) 的 LLPS 的分子机制。我们的结果表明，TDP-43 LCD 是固有无序的，具有与先前核磁共振研究一致的螺旋结构。通过模拟 TDP-43 LCD 和四个突变体的二聚化来评估相分离倾向，结果表明 A321G、W334G 和 M337V 抑制了自组装，而 G335D 促进了自组装，这与实验报道完全一致。在二聚化过程中，两个肽经历了弹性和非弹性碰撞，自缔合二聚体具有高接触和低接触状态。这些结果表明，TDP-43 LCD 的二聚化过程是动态和异质的。此外，我们在 N 端、中央区和 C 端确定了包含疏水区簇和芳香族残基的关键区域，这些区域对于 TDP-43 LCD 的自组装是必不可少的。这些具有高结合亲和力的残基可以作为接头，在 LLPS 中形成肽网络。总的来说，我们的模拟提供了驱动 TDP-43 LCD 相分离的分子间相互作用的全面图景，为生理功能和病理机制提供了深入的了解。
