De novo3D models of SARS-CoV-2 RNA elements from consensus experimental secondary structures

Abstract

The rapid spread of COVID-19 is motivating development of antivirals targeting conserved SARS-CoV-2 molecular machinery. The SARS-CoV-2 genome includes conserved RNA elements that offer potential small-molecule drug targets, but most of their 3D structures have not been experimentally characterized. Here, we provide a compilation of chemical mapping data from our and other labs, secondary structure models, and 3D model ensembles based on Rosetta's FARFAR2 algorithm for SARS-CoV-2 RNA regions including the individual stems SL1-8 in the extended 5′ UTR; the reverse complement of the 5′ UTR SL1-4; the frameshift stimulating element (FSE); and the extended pseudoknot, hypervariable region, and s2m of the 3′ UTR. For eleven of these elements (the stems in SL1–8, reverse complement of SL1–4, FSE, s2m and 3′ UTR pseudoknot), modeling convergence supports the accuracy of predicted low energy states; subsequent cryo-EM characterization of the FSE confirms modeling accuracy. To aid efforts to discover small molecule RNA binders guided by computational models, we provide a second set of similarly prepared models for RNA riboswitches that bind small molecules. Both datasets (‘FARFAR2-SARS-CoV-2’, https://github.com/DasLab/FARFAR2-SARS-CoV-2; and ‘FARFAR2-Apo-Riboswitch’, at https://github.com/DasLab/FARFAR2-Apo-Riboswitch’) include up to 400 models for each RNA element, which may facilitate drug discovery approaches targeting dynamic ensembles of RNA molecules.

To Knot or Not to Knot: Multiple Conformations of the SARS-CoV-2 Frameshifting RNA Element

“…Our reactivity data coupled with modeling show that a switch between 144 and 156 nt leads to a new peak that corresponds to the 2_2 stem-loop motif with AS1 ( Figure S14 ). Though so far not associated with a stable 3D structure 60 (unlike 3_6, 3_3, and 3_5), this alternative state may be in competition with other FSE forms. Our MSA indicates that this AS1 may be Sarbecovirus-specific, and its covariation evidence is weaker than Stem 1 ( Figure 2 ).…”

To knot or not to knot: Multiple conformations of the SARS-CoV-2 frameshifting RNA element

“…While this unknotted 2_2 can be partially explained by three groups using 2D prediction programs that do not handle pseudoknots, 20,33,34 the authors attribute this to the longer (genome-wide) sequence and the dif ferences caused by in vivo vs. in vitro experiments. 30,31 However, 3D models for these 2_2 conformations do not yield well-defined 3D structures, 60 and a 3D structure built from the 88 nt 2_2 actually recovers the 3_6 pseudoknot. 31 Combined with its weaker covariation evidence than Stem 1 (Fig.…”

Characterization of SARS2 Nsp15 Nuclease Activity Reveals it’s Mad About U

“…SARS-CoV-1 Nsp15 does not cleave 2¢methylated RNA substrate efficiently and preferentially cleaves unpaired U's within a structured RNA substrate (36). Recent papers have mapped the secondary structure of the SARS-CoV-2 genome and found evidence for highly stable RNA folding throughout the genome (45)(46)(47)(48)(49). Beyond RNA modification and secondary structure, another potential mechanism of Nsp15 nuclease regulation is through compartmentalization.…”