Cryo-EM reveals remodeling of a tandem riboswitch at 2.9 Å resolution

Cryo-EM reveals remodeling of a tandem riboswitch at 2.9 Å resolution | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article Cryo-EM reveals remodeling of a tandem riboswitch at 2.9 Å resolution Karissa Sanbonmatsu, Nathan Jespersen, Jigneshkumar Prajapati, and 1 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6422592/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted You are reading this latest preprint version Abstract Riboswitches are non-coding RNA sequences that control cellular processes through ligand binding. Conformational heterogeneity is fundamental to riboswitch functionality, yet this same attribute makes structural characterization of these mRNA elements challenging. Here, we use a combination of molecular dynamics and cryo-electron microscopy to expound the flexible nature of the glycine riboswitch tandem aptamers and characterize different structural populations. We find that Mg2+ partially stabilizes the fully folded state, resulting in one-third of the particles adopting a unique “walking man” conformation, consisting of a rigidified core and two dynamic helices, and two-thirds adopting distinct, partially folded states. Glycine interactions double the relative population of fully folded particles by stabilizing a conserved inter-aptamer Hoogsteen base pair, enabling our capture of a 2.9 Å structure for this RNA-only system. The population data show that glycine and Mg²⁺ operate synergistically: glycine enhances Mg²⁺ occupancy, while Mg2+ drives glycine specificity. Our findings indicate that cryo-electron microscopy offers a promising avenue to characterize RNA folding ensembles. Biological sciences/Molecular biology/Riboswitches Biological sciences/Biochemistry/RNA Biological sciences/Structural biology/Molecular modelling Biological sciences/Biological techniques/Structure determination/Electron microscopy/Cryoelectron microscopy Biological sciences/Genetics/Genome/Transcriptional regulatory elements Full Text Additional Declarations There is NO Competing Interest. Supplementary Files Movie1.mp4 Movie 1 Movie2.mp4 Movie 2 Movie3.mpg Movie 3 Movie4.mpg Movie 4 Cite Share Download PDF Status: Under Review Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-6422592","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":444485635,"identity":"f9c9a630-c751-4c41-b211-16af447d76e6","order_by":0,"name":"Karissa 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