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TitleCryo-electron Microscopy and Exploratory Antisense Targeting of the 28-kDa Frameshift Stimulation Element from the SARS-CoV-2 RNA Genome.
Journal, issue, pagesbioRxiv, Year 2020
Publish dateJul 20, 2020
AuthorsKaiming Zhang / Ivan N Zheludev / Rachel J Hagey / Marie Teng-Pei Wu / Raphael Haslecker / Yixuan J Hou / Rachael Kretsch / Grigore D Pintilie / Ramya Rangan / Wipapat Kladwang / Shanshan Li / Edward A Pham / Claire Bernardin-Souibgui / Ralph S Baric / Timothy P Sheahan / Victoria D Souza / Jeffrey S Glenn / Wah Chiu / Rhiju Das
PubMed AbstractDrug discovery campaigns against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) are beginning to target the viral RNA genome . The frameshift stimulation element (FSE) of the SARS-CoV- ...Drug discovery campaigns against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) are beginning to target the viral RNA genome . The frameshift stimulation element (FSE) of the SARS-CoV-2 genome is required for balanced expression of essential viral proteins and is highly conserved, making it a potential candidate for antiviral targeting by small molecules and oligonucleotides . To aid global efforts focusing on SARS-CoV-2 frameshifting, we report exploratory results from frameshifting and cellular replication experiments with locked nucleic acid (LNA) antisense oligonucleotides (ASOs), which support the FSE as a therapeutic target but highlight difficulties in achieving strong inactivation. To understand current limitations, we applied cryogenic electron microscopy (cryo-EM) and the Ribosolve pipeline to determine a three-dimensional structure of the SARS-CoV-2 FSE, validated through an RNA nanostructure tagging method. This is the smallest macromolecule (88 nt; 28 kDa) resolved by single-particle cryo-EM at subnanometer resolution to date. The tertiary structure model, defined to an estimated accuracy of 5.9 Å, presents a topologically complex fold in which the 5' end threads through a ring formed inside a three-stem pseudoknot. Our results suggest an updated model for SARS-CoV-2 frameshifting as well as binding sites that may be targeted by next generation ASOs and small molecules.
External linksbioRxiv / PubMed:32743589 / PubMed Central
MethodsEM (single particle)
Resolution6.4 - 6.9 Å
Structure data

22296

6xrz

EMDB-22296, PDB-6xrz:
The 28-kDa Frameshift Stimulation Element from the SARS-CoV-2 RNA Genome
Method: EM (single particle) / Resolution: 6.9 Å

EMDB-22297:
Nanostructure of Frameshift Stimulation Element Tagged by ATP-TTR3
Method: EM (single particle) / Resolution: 6.4 Å

Source
  • severe acute respiratory syndrome coronavirus 2
KeywordsRNA / Frameshift Stimulation Element / SARS-CoV-2 / COVID-19

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