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In vivo-like nearest neighbor parameters improve prediction of fractional RNA base-pairing in cells
We conducted a thermodynamic analysis of RNA stability in Eco80 artificial cytoplasm, which mimics in vivo conditions, and compared it to transcriptome-wide probing of mRNA. Eco80 contains 80% of Escherichia coli metabolites, with biological concentrations of metal ions, including 2 mM free Mg(2+) a...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Oxford University Press
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10639048/ https://www.ncbi.nlm.nih.gov/pubmed/37855684 http://dx.doi.org/10.1093/nar/gkad807 |
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author | Sieg, Jacob P Jolley, Elizabeth A Huot, Melanie J Babitzke, Paul Bevilacqua, Philip C |
author_facet | Sieg, Jacob P Jolley, Elizabeth A Huot, Melanie J Babitzke, Paul Bevilacqua, Philip C |
author_sort | Sieg, Jacob P |
collection | PubMed |
description | We conducted a thermodynamic analysis of RNA stability in Eco80 artificial cytoplasm, which mimics in vivo conditions, and compared it to transcriptome-wide probing of mRNA. Eco80 contains 80% of Escherichia coli metabolites, with biological concentrations of metal ions, including 2 mM free Mg(2+) and 29 mM metabolite-chelated Mg(2+). Fluorescence-detected binding isotherms (FDBI) were used to conduct a thermodynamic analysis of 24 RNA helices and found that these helices, which have an average stability of –12.3 kcal/mol, are less stable by ΔΔG(o)(37) ∼1 kcal/mol. The FDBI data was used to determine a set of Watson–Crick free energy nearest neighbor parameters (NNPs), which revealed that Eco80 reduces the stability of three NNPs. This information was used to adjust the NN model using the RNAstructure package. The in vivo-like adjustments have minimal effects on the prediction of RNA secondary structures determined in vitro and in silico, but markedly improve prediction of fractional RNA base pairing in E. coli, as benchmarked with our in vivo DMS and EDC RNA chemical probing data. In summary, our thermodynamic and chemical probing analyses of RNA helices indicate that RNA secondary structures are less stable in cells than in artificially stable in vitro buffer conditions. |
format | Online Article Text |
id | pubmed-10639048 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | Oxford University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-106390482023-11-15 In vivo-like nearest neighbor parameters improve prediction of fractional RNA base-pairing in cells Sieg, Jacob P Jolley, Elizabeth A Huot, Melanie J Babitzke, Paul Bevilacqua, Philip C Nucleic Acids Res RNA and RNA-protein complexes We conducted a thermodynamic analysis of RNA stability in Eco80 artificial cytoplasm, which mimics in vivo conditions, and compared it to transcriptome-wide probing of mRNA. Eco80 contains 80% of Escherichia coli metabolites, with biological concentrations of metal ions, including 2 mM free Mg(2+) and 29 mM metabolite-chelated Mg(2+). Fluorescence-detected binding isotherms (FDBI) were used to conduct a thermodynamic analysis of 24 RNA helices and found that these helices, which have an average stability of –12.3 kcal/mol, are less stable by ΔΔG(o)(37) ∼1 kcal/mol. The FDBI data was used to determine a set of Watson–Crick free energy nearest neighbor parameters (NNPs), which revealed that Eco80 reduces the stability of three NNPs. This information was used to adjust the NN model using the RNAstructure package. The in vivo-like adjustments have minimal effects on the prediction of RNA secondary structures determined in vitro and in silico, but markedly improve prediction of fractional RNA base pairing in E. coli, as benchmarked with our in vivo DMS and EDC RNA chemical probing data. In summary, our thermodynamic and chemical probing analyses of RNA helices indicate that RNA secondary structures are less stable in cells than in artificially stable in vitro buffer conditions. Oxford University Press 2023-10-19 /pmc/articles/PMC10639048/ /pubmed/37855684 http://dx.doi.org/10.1093/nar/gkad807 Text en © The Author(s) 2023. Published by Oxford University Press on behalf of Nucleic Acids Research. https://creativecommons.org/licenses/by-nc/4.0/This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial License (https://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com |
spellingShingle | RNA and RNA-protein complexes Sieg, Jacob P Jolley, Elizabeth A Huot, Melanie J Babitzke, Paul Bevilacqua, Philip C In vivo-like nearest neighbor parameters improve prediction of fractional RNA base-pairing in cells |
title |
In vivo-like nearest neighbor parameters improve prediction of fractional RNA base-pairing in cells |
title_full |
In vivo-like nearest neighbor parameters improve prediction of fractional RNA base-pairing in cells |
title_fullStr |
In vivo-like nearest neighbor parameters improve prediction of fractional RNA base-pairing in cells |
title_full_unstemmed |
In vivo-like nearest neighbor parameters improve prediction of fractional RNA base-pairing in cells |
title_short |
In vivo-like nearest neighbor parameters improve prediction of fractional RNA base-pairing in cells |
title_sort | in vivo-like nearest neighbor parameters improve prediction of fractional rna base-pairing in cells |
topic | RNA and RNA-protein complexes |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10639048/ https://www.ncbi.nlm.nih.gov/pubmed/37855684 http://dx.doi.org/10.1093/nar/gkad807 |
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