New insights into the function of Fascin in actin bundling: A combined theoretical and experimental study
New insights into the function of Fascin in actin bundling: A combined theoretical and experimental study
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关于肌成束蛋白在肌动蛋白捆绑中的功能的新见解：理论与实验相结合的研究
DOI：
10.1016/j.biocel.2021.106056
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Geng Dong
中科院分区：
文献类型：
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作者：
Xiaodong Wu;Bing Wen;Lirui Lin;Wenqi Shi;Dajia Li;Yinwei Cheng;Li-Yan Xu;En-Min Li;Geng Dong
Fascin是一种肌动蛋白成束蛋白，在肌动蛋白丝（F-actin）的交联中起重要作用。Fascin的磷酸化是影响其结构和功能的重要翻译后修饰。例如，Fascin-S39 D的磷酸化模拟突变显著降低了其与F-肌动蛋白的捆绑能力。本文采用分子动力学模拟和生物化学方法对Fascin的肌动蛋白成束活性进行了研究。通过MD模拟模拟从丝氨酸/苏氨酸到天冬氨酸的所有单位点突变。对于5个突变体（S146 D、S156 D、S218 D、T239 D和S259 D），发现Fascin结构域2中的突变残基与相邻残基形成盐桥相互作用，表明这些残基的突变可能潜在地降低肌动蛋白捆绑活性。此外，F-actin-bundling测定和免疫荧光技术显示S146 D和T239 D对Fascin与F-actin的bundling具有强烈的影响。最后，我们发现，单位点突变不会改变Fascin的一般形状，但Fascin-S146 D和T239 D中突变残基附近的局部结构变得不稳定，从而影响Fascin与F-actin结合的能力。这些发现表明靶向Fascin的结构域2对于药物设计将是非常有用的。此外，我们的研究表明，分子动力学模拟是一个有用的方法来筛选Fascin上的哪些残基是重要的。
Fascin, one of actin bundling proteins, plays an important role in the cross-linking of actin filaments (F-actin). Phosphorylation of Fascin is an important posttranslational modification to affect its structure and function. For example, a phosphomimetic mutation of Fascin-S39D decrease its bundling ability with F-actin significantly. In this paper, we studied the actin-bundling activity of Fascin by using molecular dynamics (MD) simulations and biochemical methods. All single-site mutations from serine/threonine to aspartic acid were mimicked by MD simulations. For five mutants (S146D, S156D, S218D, T239D and S259D), the mutated residues in domain 2 of Fascin were found to form salt-bridge interactions with an adjacent residue, indicating that mutations of these residues could potentially reduce actin-bundling activity. Further, F-actin-bundling assays and immunofluorescence technique showed S146D and T239D to have a strong effect on Fascin bundling with F-actin. Finally, we show that single-site mutations do not change the general shape of Fascin, but local structures near the mutated residues in Fascin-S146D and T239D become unstable, thereby affecting the ability of Fascin to bind with F-actin. These findings suggest that targeting domain 2 of Fascin would be very useful for the drug design. In addition, our study indicates that MD simulation is a useful method to screening which residues on Fascin are important.