A pseudouridine synthase shapes tRNA structural dynamics through both catalysis and remodeling

A pseudouridine synthase shapes tRNA structural dynamics through both catalysis and remodeling | 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 A pseudouridine synthase shapes tRNA structural dynamics through both catalysis and remodeling Julia Widom, Emily Dennis, Nico Conoan Nieves, Abigail Vaaler, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6597546/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 Transfer RNA has long served as an exemplar of a thermodynamically stable, structured RNA. Yet it undergoes significant structural changes upon binding and catalysis by diverse modification enzymes. We leveraged optical binding assays and single-molecule FRET to observe tRNA structural dynamics before and upon interaction with the conserved pseudouridine synthase Pus4/TruB. We show that unmodified and pseudouridylated tRNA similarly sample one open and two closed conformations, dynamically. Binding by Pus4 to unmodified tRNA populates additional conformational states, gradually approaching an ensemble that is adopted sooner by tRNA that was pseudouridylated prior to engaging Pus4. A catalytically incompetent mutant of Pus4 binds more slowly and remodels unmodified and pre-modified tRNAs into different conformational ensembles than wild-type enzyme. Thus, Pus4 both catalyzes a lasting chemical change on tRNA and remodels it over time, perhaps to advance subsequent steps of tRNA maturation. Biological sciences/Biophysics/Single-molecule biophysics Biological sciences/Molecular biology/Single-molecule biophysics Biological sciences/Molecular biology/RNA metabolism/RNA modification Biological sciences/Biochemistry/Proteins/RNA-binding proteins Full Text Additional Declarations There is NO Competing Interest. Supplementary Files DennisXConoanNieves2025MainTextFINAL.pdf 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. We do this by developing innovative software and high quality services for the global research community. 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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-6597546","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":457211655,"identity":"b94d84d9-55be-443a-9789-9f543367d308","order_by":0,"name":"Julia 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