Using All-Atom Potentials to Refine RNA Structure Predictions of SARS-CoV-2 Stem Loops.

Christina Bergonzo,Andrea L Szakal

doi:10.3390/ijms21176188

Christina Bergonzo, Andrea L Szakal

Open Access

https://doi.org/10.3390/ijms21176188

Copy DOI

Abstract

A considerable amount of rapid-paced research is underway to combat the SARS-CoV-2 pandemic. In this work, we assess the 3D structure of the 5′ untranslated region of its RNA, in the hopes that stable secondary structures can be targeted, interrupted, or otherwise measured. To this end, we have combined molecular dynamics simulations with previous Nuclear Magnetic Resonance measurements for stem loop 2 of SARS-CoV-1 to refine 3D structure predictions of that stem loop. We find that relatively short sampling times allow for loop rearrangement from predicted structures determined in absence of water or ions, to structures better aligned with experimental data. We then use molecular dynamics to predict the refined structure of the transcription regulatory leader sequence (TRS-L) region which includes stem loop 3, and show that arrangement of the loop around exchangeable monovalent potassium can interpret the conformational equilibrium determined by in-cell dimethyl sulfate (DMS) data.

Highlights

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a betacoronavirus which emerged in December 2019, is an ongoing pandemic [1]
Stem loops 2 and 3 (SL2 and SL3, respectively) higher order structures are of interest, since SL2 structure is conserved throughout coronaviruses and SL3 contains the conserved transcription regulatory leader sequence (TRS-L), involved in discontinuous transcription [6]
To understand the deviations in structures that arise from different treatments of an ribonucleic acid (RNA) (FARFAR2 homology modeling and structure prediction using fragment assembly, vacuo modeling using database potential, and molecular dynamics using all-atom potentials including explicit solvent), we took the representative cluster centers from each of the ten reported SARS-CoV-2 SL2 FARFAR2 clusters, and the top ten lowest energy structures from the SARS-CoV-1 NMR ensemble [20], standardized the sequence by removing the 3 A overhang, and solvated them to run molecular dynamics using modern force fields

Summary

Introduction

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a betacoronavirus which emerged in December 2019, is an ongoing pandemic [1]. To aid in quantification of SARS-CoV-2 viral RNA, the National Institute of Standards and Technology (NIST) has released research grade test material containing synthetic RNA for use in development and validation of reverse transcription polymerase chain reaction (RT-PCR) based diagnostic assays for SARS-CoV-2 [3,4]. To this end, fragment selection for quantification or for targeting could be informed by prediction of RNA higher order structure, which could impact the reverse transcription step of the assay. Stem loops 2 and 3 (SL2 and SL3, respectively) higher order structures are of interest, since SL2 structure is conserved throughout coronaviruses and SL3 contains the conserved transcription regulatory leader sequence (TRS-L), involved in discontinuous transcription [6]

Methods

Results

Discussion

Conclusion