biostudies-literaturePosani ENIGMS NIH HHS4549https://www.ebi.ac.uk/biostudies/studies/S-EPMC12084557biostudies-literatureUnknown16(1)The advent of single-particle cryogenic electron microscopy (cryo-EM) has enabled near-atomic resolution imaging of large macromolecules, enhancing functional insights. However, current cryo-EM refinement tools condense all single-particle images into a single structure, which can misrepresent highly flexible molecules like RNAs. Here, we combine molecular dynamics simulations with cryo-EM density maps to better account for the structural dynamics of a complex and biologically relevant RNA macromolecule. Namely, using metainference, a Bayesian method, we reconstruct an ensemble of structures of the group II intron ribozyme, which better matches experimental data, and we reveal inaccuracies of single-structure approaches in modeling flexible regions. An analysis of all RNA-containing structNature communicationsEnsemble refinement of mismodeled cryo-EM RNA structures using all-atom simulations.PMC12084557R35 GM141706Bussi GBonomi MPosani EMagistrato AHaack DToor NJanos PfalseEnsemble refinement of mismodeled cryo-EM RNA structures using all-atom simulations.The advent of single-particle cryogenic electron microscopy (cryo-EM) has enabled near-atomic resolution imaging of large macromolecules, enhancing functional insights. However, current cryo-EM refinement tools condense all single-particle images into a single structure, which can misrepresent highly flexible molecules like RNAs. Here, we combine molecular dynamics simulations with cryo-EM density maps to better account for the structural dynamics of a complex and biologically relevant RNA macromolecule. Namely, using metainference, a Bayesian method, we reconstruct an ensemble of structures of the group II intron ribozyme, which better matches experimental data, and we reveal inaccuracies of single-structure approaches in modeling flexible regions. An analysis of all RNA-containing struct2025-01-01T00:00:00Z2025 May2026-04-08T19:53:35.022Z2026-04-08T14:32:01.903ZS-EPMC120845574037969910.1038/s41467-025-59769-0