De novo designed bifunctional proteins for targeted protein degradation

De novo designed bifunctional proteins for targeted protein degradation | 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 De novo designed bifunctional proteins for targeted protein degradation Derek Woolfson, Bram Mylemans, Boguslawa Korona, Amanda Acevedo-Jake, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8425103/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 Targeted protein degradation (TPD) is a therapeutic strategy to remove disease-causing proteins by routing them to the ubiquitin-proteasome, autophagy, or lysosme machineries. For instance, proteolysis-targeting chimeras (PROTACs) are synthetic hetero-bifunctional small molecules that simultaneously bind the target and an E3 ubiquitin ligase to drive ubiquitination and degradation by the proteasome. Despite considerable success, designing such molecules is challenging and the number of currently addressable ubiquitin E3 ligases is limited. Here we demonstrate hetero-bifunctional de novo designed proteins as alternatives for TPD to access more targets and ligases. First, we develop a stable and highly adaptable helix-turn-helix scaffold for presenting different binding sites. Next, we use computational protein design to incorporate and embellish hot-spot- binding sites to target BCL-xL, plus short linear motifs (SLiMs) for KLHL20 ligase recruitment. The resulting mono- and bi-functionalised proteins bind the targets in vitro, and the latter degrade BCL-xL in cells leading to apoptosis. Biological sciences/Chemical biology/Protein design Biological sciences/Cell biology/Proteolysis/Protein quality control Full Text Additional Declarations There is NO Competing Interest. Supplementary Files MylemansPROTACCNatBiotechDec2025SIsubmitted.pdf Mylemans_PROTACC_NatBiotech_Dec2025_SI_submitted 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-8425103","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":571959191,"identity":"2899ec30-a980-47c1-af85-d321e400112d","order_by":0,"name":"Derek Woolfson","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABEUlEQVRIie2RsWrDMBCGzwic5RKvDg7xK8gUDIE2fhWFgLvEEOiSoYPB4KnJHij0FTpmvGLQpNI3CIW8QLoUOrRUciklEIWMHfRNEqcP/XcH4HD8QzJAzhBg0Cn17a0EfQaP2hoeV5Ky1yqI+pm3NooPcFLhFPwprGu+Ok9ZXCIGjdxdbbaD+KEigtsxcEUWpfvIZipHDPPri0LdIJe+IJBT4M+lXSnqBrMQ06ioBXIfOYFPwF9swXDfKhioNBppJa6N8nVSmbNPo8AsjTytgNSKV5M1WFLhvLn/6SXvL5UOJnNBk9UU+5b2s85dstMTG5qJhR8bkcVV8/S6fx8Pe0ocT8Z+t3CAsG7F4XA4HOfwDYcJWO161LX0AAAAAElFTkSuQmCC","orcid":"https://orcid.org/0000-0002-0394-3202","institution":"University of Bristol","correspondingAuthor":true,"prefix":"","firstName":"Derek","middleName":"","lastName":"Woolfson","suffix":""},{"id":571959192,"identity":"fc1ef0ad-424b-495a-a06a-288d80162c7b","order_by":1,"name":"Bram Mylemans","email":"","orcid":"","institution":"School of Chemistry, University of Bristol, Bristol, UK; 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