Structural insight into receptor and substrate specificity of Cytotoxic Necrotizing Factor CNF1 from Escherichia coli

Structural insight into receptor and substrate specificity of Cytotoxic Necrotizing Factor CNF1 from Escherichia coli | 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 Structural insight into receptor and substrate specificity of Cytotoxic Necrotizing Factor CNF1 from Escherichia coli Wulf Blankenfeldt, Shuangshuang Dong, Peer Lukat, Stefan Schmelz, and 4 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-9579774/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 cytotoxic necrotizing factors (CNFs) are bacterial toxins that deamidate or transaminate a conserved glutamine in switch-II of RhoGTPases, leading to their constitutive activation and host cell death. Despite early discovery in the 1980s, structural insight into the mechanisms underlying CNF activity is limited. Here, we report crystal structures of Escherichia coli CNF1 alone and in complex with its cellular receptor Lu/BCAM and with its substrate RhoA. The overall structure of CNF1 differs from the Yersinia pseudotuberculosis homologue CNFY, yet individual domains are well conserved, revealing a modular architecture with flexible interdomain organization, supported by small-angle X-ray scattering (SAXS). Lu/BCAM binds to a different CNF1 segment than previously reported, and the structure explains why CNFY does not recognize this receptor. Interaction with the toxin refolds both switch regions in RhoA, leading to disorder of switch-I but specific recognition of the switch-II sequence, hence explaining the GTPase-selectivity of CNFs. Biological sciences/Structural biology/X-ray crystallography Biological sciences/Microbiology/Pathogens Biological sciences/Biochemistry/Proteins Biological sciences/Structural biology/SAXS Full Text Additional Declarations There is NO Competing Interest. Supplementary Files cnf1supplementaryinformation20260429.pdf Supplementary information for "Structural insight into receptor and substrate specificity of Cytotoxic Necrotizing Factor CNF1 from Escherichia coli" 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-9579774","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":639629787,"identity":"41ae48d0-6851-4979-af84-9509cc3f3c2b","order_by":0,"name":"Wulf 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