Mechanisms of SecM-Mediated Stalling in the Ribosome
Mechanisms of SecM-Mediated Stalling in the Ribosome
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DOI：
10.1016/j.bpj.2012.06.005
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发表时间：
2012-07-18
影响因子：
3.4
通讯作者：
Schulten, Klaus
中科院分区：
文献类型：
--
作者：
Gumbart, James;Schreiner, Eduard;Schulten, Klaus
翻译的新生肽调节是基因表达的常见调节机制。在这种机制中，当新生肽仍在核糖体的出口通道中时，它会诱导翻译暂停，从而控制下游基因的表达。 SecM 就是一个例子，它在自身翻译过程中抑制核糖体肽基转移酶中心 (PTC) 中肽键的形成，从而上调蛋白质转位酶 SecA 的表达。尽管生化实验和冷冻电镜数据已经鉴定出一些参与 SecM 识别的残基，但通过核糖体将 SecM 连接到 PTC 的相互作用残基的完整途径尚未最终确定。在这里，利用冷冻电子显微镜数据，我们通过分子动力学灵活拟合导出了第一个（据我们所知）SecM 停滞核糖体的原子模型，并配有 P 位点和 A 位点 tRNA。随后，我们对天然和突变的 SecM 停滞核糖体进行了模拟，以研究关键 SecM 残基 R163 和 PTC 之间可能的相互作用途径。特别是，模拟揭示了 SecM 在改变核糖体中 tRNA 位置方面的作用，从而证明了出口隧道中 SecM 的存在如何引起失速。最后，引导分子动力学模拟（其中 SecM 被拉向隧道出口）表明 SecA 从核糖体外部与 SecM 相互作用如何缓解失速。
Nascent-peptide modulation of translation is a common regulatory mechanism of gene expression. In this mechanism, while the nascent peptide is still in the exit tunnel of the ribosome, it induces translational pausing, thereby controlling the expression of downstream genes. One example is SecM, which inhibits peptide-bond formation in the ribosome's peptidyl transferase center (PTC) during its own translation, upregulating the expression of the protein translocase SecA. Although biochemical experiments and cryo-electron microscopy data have led to the identification of some residues involved in SecM recognition, the full pathway of interacting residues that connect SecM to the PTC through the ribosome has not yet been conclusively established. Here, using the cryo-electron microscopy data, we derived the first (to our knowledge) atomic model of the SecM-stalled ribosome via molecular-dynamics flexible fitting, complete with P-and A-site tRNAs. Subsequently, we carried out simulations of native and mutated SecM-stalled ribosomes to investigate possible interaction pathways between a critical SecM residue, R163, and the PTC. In particular, the simulations reveal the role of SecM in altering the position of the tRNAs in the ribosome, and thus demonstrate how the presence of SecM in the exit tunnel induces stalling. Finally, steered molecular-dynamics simulations in which SecM was pulled toward the tunnel exit suggest how SecA interacting with SecM from outside the ribosome relieves stalling.