Synthetic yeast genome SCRaMbLEing uncovers a new role for ribosomal proteins in genetic code expansion

Synthetic yeast genome SCRaMbLEing uncovers a new role for ribosomal proteins in genetic code expansion | 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 Synthetic yeast genome SCRaMbLEing uncovers a new role for ribosomal proteins in genetic code expansion Yizhi Cai, James Sanders, Stefan Hoffmann, Ewan Moody, Mark McCullough This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7505907/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 The translation of proteins with non-canonical amino acids (ncAAs) has emerged as a powerful technology for embedding new functional elements into proteins, enabling the development of novel enzymes, materials, and biopharmaceuticals. However, the utility of this approach has been hindered by weak translation efficiencies. To address this challenge, we sought to substantially improve orthogonal translation in Saccharomyces cerevisiae. We first evaluated recently described ∆NPylRS-class pyrrolysyl-tRNA synthetase systems and identified a homolog with∼5.4-fold higher activity than the best previously reported pyrrolysyl system. Building on this advance, we leveraged the SCRaMbLE system in the semi-synthetic yeast strain Syn6.5 to generate structural genomic variation, and identified strains with enhanced ncAA incorporation. Pooling genomic alterations across enhanced strains, we identified an association for the deletion of several ribosomal protein genes with the increased production of ncAA-containing protein. These findings demonstrate a previously unrecognized role for ribosomal proteins in enabling alternate genetic codes. Biological sciences/Systems biology/Synthetic biology Biological sciences/Chemical biology/Chemical genetics Full Text Additional Declarations There is NO Competing Interest. Supplementary Files SI1uragrowthassay.png Figure S1: Characterization of the URA3 Growth Assay. SI2MSdata.png Figure S2: Mass Spectra of msGFP Protein 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-7505907","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":532594328,"identity":"4245d4f9-f66c-45d2-b504-e2a4493d8189","order_by":0,"name":"Yizhi 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