Structure and function of the EDEM:PDI ERAD checkpoint complex

Structure and function of the EDEM:PDI ERAD checkpoint complex | 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 and function of the EDEM:PDI ERAD checkpoint complex Pietro Roversi, Charlie Hitchman, Andrea Lia, Gabriela Chiritoiu, and 15 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6066341/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 ERAD glycoprotein misfolding checkpoint complex de-mannosylates misfolded glycoproteins targeting them to retrotranslocation, ubiquitination, and proteasomal degradation. The complex comprises an Endoplasmic Reticulum-Degradation Enhancing α-Mannosidase (EDEM) and a Protein Disulfide Isomerase (PDI). We solved Cryo-EM structures of the Chaetomium thermophilum (Ct) EDEM:PDI complex, both by itself and in complex with a classic ERAD substrate, α1-antitrypsin (A1AT-NHK). The EDEM catalytic domain nestles within the PDI arc, while A1AT-NHK binds EDEM’s C-terminal flexible domains. Mass spectrometry reveals a disulfide bond between A1AT-NHK and an exposed Cys in the protease-associated domain of EDEM. Non-reducing SDS-PAGE analysis of protein samples from mammalian cells co-transfected with EDEM:PDI and A1AT-NHK show a shift of the EDEM:PDI band to higher molecular weight. Redox chemistry between EDEM and PDI disulfide bonds generates oxidized, demannosylation-competent EDEM and reduced PDI and primes the PDI to function as the ERAD reductase, facilitating client retrotranslocation. Biological sciences/Structural biology/Electron microscopy/Cryoelectron microscopy Biological sciences/Biochemistry/Glycobiology ERAD EDEM PDI CryoEM A1AT-NHK EDEM3 ERp46 Full Text Additional Declarations There is NO Competing Interest. Supplementary Files EDEMPDI19Feb2025SI.pdf SupplementaryVideo1.mp4 Supplementary Video 1 SupplementaryVideo2.mp4 Supplementary Video 2 SupplementaryVideo3.mp4 Supplementary Video 3 SupplementaryVideo4.mp4 Supplementary Video 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. 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-6066341","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":425672501,"identity":"50aafdf2-0bce-428b-8da1-06e73a86258f","order_by":0,"name":"Pietro 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