Protein unties the pseudoknot: S1-mediated unfolding of RNA higher order structure
Protein unties the pseudoknot: S1-mediated unfolding of RNA higher order structure
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DOI：
10.1093/nar/gkz1166
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发表时间：
2020-02-28
影响因子：
14.9
通讯作者：
Walter, Nils G.
中科院分区：
文献类型：
--
作者：
Lund, Paul E.;Chatterjee, Surajit;Walter, Nils G.
除了核糖体以外的各种细胞过程之外，核糖体蛋白 Si 在许多大肠杆菌 mRNA 的翻译起始步骤中发挥着重要作用，特别是那些具有弱 Shine-Dalgarno 序列或结构化 5' UTR 的mRNA。在所有情况下，Si 的 RNA 结合活性是其功能的核心特征。虽然Si亲和力的序列决定因素以及Si与简单二级结构相互作用的许多要素是已知的，但蛋白质与更复杂的二级和三级结构的RNA相互作用的机制细节却知之甚少。在这里，我们研究了 Si 与 I 类翻译 preQ(1) 核糖开关的充分表征的 H 型假结的相互作用，作为高度结构化的 RNA 模型，其构象和结构动力学可以通过添加不同结合亲和力的配体（特别是 preQ(1)、鸟嘌呤和 2,6-二氨基嘌呤）来调整。结合生化和单分子荧光方法，我们发现 Si 优先与折叠较少的假结相互作用，并促进动态的、部分展开的构象。 Si解折叠RNA的能力与假结的结构稳定性成反比。这些机制见解描绘了 Si 陪伴的结构化 RNA 展开的范围和局限性。
Ribosomal protein Si plays important roles in the translation initiation step of many Escherichia coli mRNAs, particularly those with weak Shine-Dalgarno sequences or structured 5' UTRs, in addition to a variety of cellular processes beyond the ribosome. In all cases, the RNA-binding activity of Si is a central feature of its function. While sequence determinants of Si affinity and many elements of the interactions of Si with simple secondary structures are known, mechanistic details of the protein's interactions with RNAs of more complex secondary and tertiary structure are less understood. Here, we investigate the interaction of Si with the well-characterized H-type pseudoknot of a class-I translational preQ(1) riboswitch as a highly structured RNA model whose conformation and structural dynamics can be tuned by the addition of ligands of varying binding affinity, particularly preQ(1), guanine, and 2,6-diaminopurine. Combining biochemical and single molecule fluorescence approaches, we show that Si preferentially interacts with the less folded form of the pseudoknot and promotes a dynamic, partially unfolded conformation. The ability of Si to unfold the RNA is inversely correlated with the structural stability of the pseudoknot. These mechanistic insights delineate the scope and limitations of Si-chaperoned unfolding of structured RNAs.