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. 2023 Apr 27;14:2416. doi: 10.1038/s41467-023-38098-0

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© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 3 — The model-predicted mRNA structures and ribosome-mRNA binding free energies for the A MS2 ON switch (M2 riboswitch) and the B MS2 OFF switch (M7o riboswitch) across its four most relevant states. State 1 shows the initial mRNA structure and its Gibbs free energy of folding (ΔG_initial) when the MS2 coat protein is not bound. State 2 shows the change in mRNA structure and the final Gibbs free energy (ΔG_final) when the ribosome binds to the mRNA in State 1. The free energy model for calculating ΔG_final includes hybridization between the mRNA and ribosomal RNA (ΔG_mRNA-rRNA), base pairing between the tRNA and start codon (ΔG_start), and energetic penalties for non-optimal spacing (ΔG_spacing) and an inaccessible standby site (ΔG_standby). State 3 shows how the mRNA structure changes when the MS2 coat protein binds to its cognate RNA aptamer with binding free energy ΔG_ligand, which results in a change in mRNA folding free energy (ΔG_initial). State 4 shows the change in mRNA structure and final Gibbs free energy (ΔG_final) when the ribosome binds to the mRNA in State 3. The translation initiation rates are predicted based on the difference in initial and final Gibbs free energies (ΔG_total) according to Boltzmann’s relationship. Nucleotides are colored according to their interactions, including (blue) the RNA aptamer domain, (red) the last 9 nucleotides of the 16S ribosomal RNA, (green) the Shine–Dalgarno sequence, (purple) the start codon, and (orange) the standby site. The brown bar is the ribosomal footprint for initiation.