The SARS-CoV-2 Programmed −1 Ribosomal Frameshifting Element Crystal Structure Solved to 2.09 Å Using Chaperone-Assisted RNA Crystallography,ACS Chemical Biology

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The SARS-CoV-2 Programmed −1 Ribosomal Frameshifting Element Crystal Structure Solved to 2.09 Å Using Chaperone-Assisted RNA Crystallography
ACS Chemical Biology ( IF 3.5 ) Pub Date : 2021-07-30 , DOI: 10.1021/acschembio.1c00324
Christina Roman ₁ , Anna Lewicka ₁ , Deepak Koirala ₂ , Nan-Sheng Li ₁ , Joseph A Piccirilli _{1,

3}

Affiliation

The programmed −1 ribosomal frameshifting element (PFSE) of SARS-CoV-2 is a well conserved structured RNA found in all coronaviruses’ genomes. By adopting a pseudoknot structure in the presence of the ribosome, the PFSE promotes a ribosomal frameshifting event near the stop codon of the first open reading frame Orf1a during translation of the polyprotein pp1a. Frameshifting results in continuation of pp1a via a new open reading frame, Orf1b, that produces the longer pp1ab polyprotein. Polyproteins pp1a and pp1ab produce nonstructural proteins NSPs 1–10 and NSPs 1–16, respectively, which contribute vital functions during the viral life cycle and must be present in the proper stoichiometry. Both drugs and sequence alterations that affect the stability of the −1 programmed ribosomal frameshifting element disrupt the stoichiometry of the NSPs produced, which compromise viral replication. For this reason, the −1 programmed frameshifting element is considered a promising drug target. Using chaperone assisted RNA crystallography, we successfully crystallized and solved the three-dimensional structure of the PFSE. We observe a three-stem H-type pseudoknot structure with the three stems stacked in a vertical orientation stabilized by two triple base pairs at the stem 1/stem 2 and stem 1/stem 3 junctions. This structure provides a new conformation of PFSE distinct from the bent conformations inferred from midresolution cryo-EM models and provides a high-resolution framework for mechanistic investigations and structure-based drug design.

中文翻译：

使用分子伴侣辅助 RNA 晶体学将 SARS-CoV-2 编程的 −1 核糖体移码元件晶体结构解析为 2.09 Å

SARS-CoV-2 的程序化 -1 核糖体移码元件 (PFSE) 是所有冠状病毒基因组中发现的高度保守的结构 RNA。通过在核糖体存在的情况下采用假结结构，PFSE 在多蛋白 pp1a 的翻译过程中促进第一个开放阅读框 Orf1a 终止密码子附近的核糖体移码事件。移码导致 pp1a 通过新的开放阅读框 Orf1b 延续，从而产生更长的 pp1ab 多蛋白。多蛋白 pp1a 和 pp1ab 分别产生非结构蛋白 NSP 1-10 和 NSP 1-16，它们在病毒生命周期中发挥重要功能，并且必须以正确的化学计量存在。影响 -1 编程核糖体移码元件稳定性的药物和序列改变都会破坏所产生的 NSP 的化学计量，从而损害病毒复制。因此，-1 编程移码元件被认为是有前途的药物靶点。利用伴侣辅助RNA晶体学，我们成功结晶并解析了PFSE的三维结构。我们观察到三茎 H 型假结结构，其中三个茎以垂直方向堆叠，由茎 1/茎 2 和茎 1/茎 3 连接处的两个三碱基对稳定。该结构提供了 PFSE 的新构象，与中分辨率冷冻电镜模型推断的弯曲构象不同，并为机理研究和基于结构的药物设计提供了高分辨率框架。

更新日期：2021-08-20

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