Cryo-electron Microscopy and Exploratory Antisense Targeting of the 28-kDa Frameshift Stimulation Element from the SARS-CoV-2 RNA Genome

BioRxiv : the Preprint Server for Biology
K. ZhangRhiju Das

Abstract

Drug discovery campaigns against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) are beginning to target the viral RNA genome 1, 2 . 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 3-6 . 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 7 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 insid...Continue Reading

Citations

Nov 10, 2020·Nucleic Acids Research·I. ManfredoniaDanny Incarnato
Jan 21, 2021·The Journal of Physical Chemistry. B·Qiyao Zhu, Tamar Schlick
Dec 29, 2020·Biochemical Society Transactions·I. Manfredonia, Danny Incarnato
Mar 31, 2021·Proceedings of the National Academy of Sciences of the United States of America·Seyed-Fakhreddin TorabiJoan A Steitz
Apr 29, 2021·Acta Pharmacologica Sinica·Peng-Xuan RenFang Bai
May 9, 2021·Nature Methods·Rhiju Das
May 27, 2021·Journal of the American Chemical Society·Sandro BottaroKresten Lindorff-Larsen
May 28, 2021·Ecotoxicology and Environmental Safety·Fan ZhangWillie J G M Peijnenburg
May 28, 2021·NAR Genomics and Bioinformatics·Ryan J AndrewsWalter N Moss

Methods Mentioned

BETA
x-ray scattering
NMR
Antisense oligonucleotides
electrophoresis
electron microscopy
transfection
Assay
transfections
PCR
in vitro transcription

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