Paloni Matteo, Bailly Rémy, Ciandrini Luca, Barducci Alessandro
Centre de Biochimie Structurale (CBS), INSERM, CNRS, Université de Montpellier, Montpellier, France.
J Phys Chem B. 2020 Oct 15;124(41):9009-9016. doi: 10.1021/acs.jpcb.0c06288. Epub 2020 Oct 5.
Membraneless organelles are dynamical cellular condensates formed via biomolecular liquid-liquid phase separation of proteins and RNA molecules. Multiple evidence suggests that in several cases disordered proteins are structural scaffolds that drive the condensation by forming a dynamic network of inter- and intramolecular contacts. Despite the blooming research activity in this field, the structural characterization of these entities is very limited, and we still do not understand how the phase behavior is encoded in the amino acid sequences of the scaffolding proteins. Here we exploited explicit-solvent atomistic simulations to investigate the N-terminal disordered region of DEAD-box helicase 4 (NDDX4), which is a well-established model for phase separation. Notably, we determined NDDX4 conformational ensemble at the single-molecule level, and we relied on a "divide-and-conquer" strategy, based on simulations of various protein fragments at high concentration, to probe intermolecular interactions in conditions mimicking real condensates. Our results provide a high-resolution picture of the molecular mechanisms underlying phase separation in agreement with NMR and mutagenesis data and suggest that clusters of arginine and aromatic residues may stabilize the assembly of several condensates.
无膜细胞器是通过蛋白质和RNA分子的生物分子液-液相分离形成的动态细胞凝聚物。多项证据表明，在某些情况下，无序蛋白质是通过形成分子间和分子内接触的动态网络来驱动凝聚的结构支架。尽管该领域的研究活动蓬勃发展，但这些实体的结构表征非常有限，我们仍然不了解相行为是如何在支架蛋白的氨基酸序列中编码的。在这里，我们利用显式溶剂原子模拟来研究DEAD盒解旋酶4的N端无序区域（NDDX4），它是一个成熟的相分离模型。值得注意的是，我们在单分子水平上确定了NDDX4的构象集合，并基于高浓度下各种蛋白质片段的模拟，采用“分而治之”策略来探测模拟真实凝聚物条件下的分子间相互作用。我们的结果提供了与核磁共振和诱变数据一致的相分离分子机制的高分辨率图景，并表明精氨酸和芳香族残基簇可能稳定几种凝聚物的组装。
