Structure of the giant RNA polymerase ejected from coliphage N4

Structure of the giant RNA polymerase ejected from coliphage N4 | 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 Structure of the giant RNA polymerase ejected from coliphage N4 Nathan Bellis, Ravi Lokareddy, Mikhail Pavlenok, Stephanie Cooper Horton, and 6 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7746245/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 Schitoviruses are widespread prokaryotic viruses that encapsidate a giant (~3,500-residue) virion-associated RNA polymerase (vRNAP). During infection, vRNAP is expelled into Gram-negative bacteria, along with two additional ejection proteins, to assemble a transient DNA-ejectosome that becomes transcriptionally active, initiating viral replication. Here, we present an integrative structural analysis of the coliphage N4 vRNAP (gp50). We find that this 383 kDa enzyme is a multi-domain, single-chain RNA polymerase, structurally distinct from both compact single-chain RNAPs and large multi-subunit holoenzymes. vRNAP is composed of loosely connected domains and exhibits an intramolecular mode of allosteric regulation through its C-terminal domain. Comparative analysis of intact and genome-released virions identified gp51, which forms an outer-membrane complex, and gp52, which assembles a periplasmic tunnel. These proteins generate heterogeneous pores that facilitate the release of vRNAP. We further uncover a signaling hub in the phage tail, composed of the receptor-binding protein, tail tube, and tail plug, that detects receptor engagement and orchestrates the release of ejection proteins. We propose that the beads-on-a-string architecture of vRNAP enables the translocation of megadalton-scale protein complexes through the ~35 Å channel formed by the tail and ejection proteins. These findings establish N4 as a distinctive model for protein translocation through biological channels. Biological sciences/Structural biology/Electron microscopy/Cryoelectron microscopy Biological sciences/Microbiology/Virology/Phage biology Full Text Additional Declarations There is NO Competing Interest. Supplementary Files nrreportingsummary.docx modelvalidationsmerged.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. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. 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-7746245","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":528620661,"identity":"b7e861a4-d4ca-417f-b48e-653380e73185","order_by":0,"name":"Nathan Bellis","email":"","orcid":"","institution":"University of Alabama at Birmingham","correspondingAuthor":false,"prefix":"","firstName":"Nathan","middleName":"","lastName":"Bellis","suffix":""},{"id":528620662,"identity":"23160e8c-8972-46e8-9977-f94a73929253","order_by":1,"name":"Ravi 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