NMR characterization and ligand binding site of the stem loop 2 motif (s2m) from the Delta variant of SARS-CoV-2
- Tobias Matzel1,
- Maria Wirtz Martin2,
- Alexander Herr1,
- Anna Wacker1,
- Christian Richter1,
- Sridhar Sreeramulu3 and
- Harald Schwalbe4,5
- 1 Goethe-University, Institute for Organic Chemistry and Chemical Biology, Center for Biomolecular Magnetic Resonance;
- 2 Goethe University, Institute for Organic Chemistry and Chemical Biology, Center for Biomolecular Magnetic Resonance;
- 3 Goethe-University,Institute for Organic Chemistry and Chemical Biology, Center for Biomolecular Magnetic Resonance;
- 4 University of Frankfurt, Institute for Organic Chemistry and Chemical Biology, Center for Biomolecular Magnetic Resonance
- ↵* Corresponding author; email: schwalbe{at}nmr.uni-frankfurt.de
Abstract
The stem loop 2 motif (s2m) in SARS-CoV-2 (SCoV-2) is located in the 3’-UTR. Although s2m has been reported to display characteristics of a mobile genomic element that might lead to an evolutionary advantage, its function has remained unknown. The secondary structure of the original SCoV-2 RNA sequence (Wuhan-Hu-1) was determined by NMR in late 2020, delineating the base pairing pattern and revealing substantial differences in secondary structure compared to SARS-CoV-1 (SCoV-1). The existence of a single G29742-A29756 mismatch in the upper stem of s2m leads to its destabilization and impedes a complete NMR analysis. With Delta, a variant of concern has evolved with one mutation compared to the original sequence that replaces G29742 by U29742. We show here that this mutation results in a more defined structure at ambient temperature accompanied by a rise in melting temperature. Consequently, we were able to identify over 90 % of the relevant NMR resonances using a combination of selective RNA labeling and filtered 2D NOESY as well as 4D NMR experiments. We present a comprehensive NMR analysis of the secondary structure, (sub-) nanosecond dynamics and ribose conformation of s2m Delta based on heteronuclear 13C NOE and T1 measurements and ribose carbon chemical shift-derived canonical coordinates. We further show that the G29742U mutation in Delta has no influence on the druggability of s2m compared to the Wuhan-Hu-1 sequence. With the assignment at hand, we identify the flexible regions of s2m as primary site for small molecule binding.
Keywords
- Received November 27, 2023.
- Accepted March 8, 2024.
- Published by Cold Spring Harbor Laboratory Press for the RNA Society
This article, published in RNA, is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0/.