Meranda M. Masse, Rachel B. Hutchinson, Christopher E. Morgan, Heather J. Allaman, Hongqing Guan, Edward W. Yu and Silvia Cavagnero*,
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引用次数: 0
Abstract
Interactions between ribosome-bound nascent chains (RNCs) and ribosomal components are critical to elucidate the mechanism of cotranslational protein folding. Nascent protein–ribosome contacts within the ribosomal exit tunnel were previously assessed mostly in the presence of C-terminal stalling sequences, yet little is known about contacts taking place in the absence of these strongly interacting motifs. Further, there is nearly no information about ribosomal proteins (r-proteins) interacting with nascent chains within the outer surface of the ribosome. Here, we combine chemical cross-linking, single-particle cryo-EM, and fluorescence anisotropy decays to determine the structural features of ribosome-bound apomyoglobin (apoMb). Within the ribosomal exit tunnel core, interactions are similar to those identified in previous reports. However, once the RNC enters the tunnel vestibule, it becomes more dynamic and interacts with ribosomal RNA (rRNA) and the L23 r-protein. Remarkably, on the outer surface of the ribosome, RNCs interact mainly with a highly conserved nonpolar patch of the L23 r-protein. RNCs also comprise a compact and dynamic N-terminal region lacking contact with the ribosome. In all, apoMb traverses the ribosome and interacts with it via its C-terminal region, while N-terminal residues sample conformational space and form a compact subdomain before the entire nascent protein sequence departs from the ribosome.
Single-particle cryogenic EM and fluorescence anisotropy decays show that a nascent globin interacts with the L23 ribosomal protein. The interaction site is highly nonpolar, suggesting a chaperone behavior.
核糖体结合新生链(RNC)与核糖体成分之间的相互作用对于阐明共翻译蛋白质折叠机制至关重要。以前对核糖体出口隧道内新生蛋白与核糖体接触情况的评估大多是在存在 C 端停滞序列的情况下进行的,但对于在没有这些强相互作用基团的情况下发生的接触却知之甚少。此外,关于核糖体蛋白(r 蛋白)与核糖体外表面新生链相互作用的信息几乎为零。在这里,我们结合化学交联、单颗粒低温电子显微镜和荧光各向异性衰减来确定核糖体结合的肌红蛋白(apomyoglobin,apoMb)的结构特征。在核糖体出口隧道核心内,相互作用与以前报告中确定的相互作用相似。然而,一旦 RNC 进入隧道前庭,它就会变得更加活跃,并与核糖体 RNA(rRNA)和 L23 r 蛋白相互作用。值得注意的是,在核糖体的外表面,RNC 主要与 L23 r 蛋白的一个高度保守的非极性斑块相互作用。RNC 还包括一个紧凑而动态的 N 端区域,该区域缺乏与核糖体的接触。总之,apoMb穿过核糖体并通过其C端区域与核糖体相互作用,而N端残基则在整个新生蛋白质序列离开核糖体之前对构象空间进行采样并形成一个紧凑的亚域。该相互作用位点高度非极性,表明其具有伴侣行为。
期刊介绍:
ACS Central Science publishes significant primary reports on research in chemistry and allied fields where chemical approaches are pivotal. As the first fully open-access journal by the American Chemical Society, it covers compelling and important contributions to the broad chemistry and scientific community. "Central science," a term popularized nearly 40 years ago, emphasizes chemistry's central role in connecting physical and life sciences, and fundamental sciences with applied disciplines like medicine and engineering. The journal focuses on exceptional quality articles, addressing advances in fundamental chemistry and interdisciplinary research.