Biocatalytic Radical C(sp³)–N Coupling via Active Site Templating

Biocatalytic Radical C(sp³)–N Coupling via Active Site Templating | 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 Biocatalytic Radical C(sp³)–N Coupling via Active Site Templating Todd Hyster, Zayed Alassad This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-9204910/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 Stereoselective nucleophilic substitution to access α tertiary amines relies on copper catalyzed radical approaches in which the substitution is mediated by metal–anilide coordination.1 These systems, however, are constrained by competing arene radical alkylation pathways.2 Here we report a distinct photoenzymatic mechanism for enantioconvergent nucleophilic substitution that operates without metal coordination to the nucleophile. Six rounds of protein engineering yielded a variant of a flavin dependent oxidoreductase that promotes C(sp³)–N coupling between tertiary alkyl halides and simple anilines in good yields, with high chemoselectivity for N over C alkylation and high enantioselectivity across a broad substrate range. Multivariate statistical analysis, density functional theory, and mechanistic experiments show that the active site templates π stacking, hydrogen bonding, and water bridged interactions between a tertiary radical and the aniline lone pair to generate an intermolecular n→SOMO hyperconjugative complex that is energetically disfavored in bulk solution, thereby simultaneously lowering the radical oxidation potential and suppressing arene addition.3 This work uncovers a previously inaccessible, copper free manifold for nucleophilic substitution at sterically congested carbon centers and expands how enzymes can catalyze C(sp³)–N bond formation with control over both stereo and chemoselectivity. Physical sciences/Chemistry/Catalysis/Biocatalysis Physical sciences/Chemistry/Photochemistry/Photocatalysis Full Text Additional Declarations There is NO Competing Interest. Supplementary Files model2026.csv Modelling Excel File RedoxCalculations.xlsx Excel of Redox Calculations SupplementalInformation.pdf Supplemental Information SIDFT.zip DFT Calculation Files Codeformultivariatemodel.docx Multivariate Model 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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