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dc.contributor.authorSeebald, Leah
dc.contributor.authorMadec, Amael
dc.contributor.authorImperiali, Barbara
dc.date.accessioned2020-07-20T21:12:32Z
dc.date.available2020-07-20T21:12:32Z
dc.date.issued2019-11
dc.identifier.issn1439-4227
dc.identifier.issn1439-7633
dc.identifier.urihttps://hdl.handle.net/1721.1/126269
dc.description.abstractHigh-throughput small-molecule screening in drug discovery processes commonly rely on fluorescence-based methods including fluorescent polarization and fluorescence/Förster resonance energy transfer. These techniques use highly accessible instrumentation; however, they can suffer from high false-negative rates and background signals, or might involve complex schemes for the introduction of fluorophore pairs. Herein we present the synthesis and application of fluorescent nucleoside analogues as the foundation for directed approaches for competitive binding analyses. The general approach describes selective fluorescent environment-sensitive (ES) nucleoside analogues that are adaptable to diverse enzymes that act on nucleoside-based substrates. We demonstrate screening a set of uridine analogues and development of an assay for fragment-based lead discovery with the TcdB glycosyltransferase (GT), an enzyme associated with virulence in Clostridium difficile. The uridine-based probe used for this high-throughput screen has a KD value of 7.2 μm with the TcdB GT and shows a >30-fold increase in fluorescence intensity upon binding. The ES-based probe assay is benchmarked against two other screening approaches.en_US
dc.description.sponsorshipNational Institutes of Health (Grants GM097241, GM131627)en_US
dc.language.isoen
dc.publisherWileyen_US
dc.relation.isversionofhttp://dx.doi.org/10.1002/cbic.201900671en_US
dc.rightsCreative Commons Attribution-Noncommercial-Share Alikeen_US
dc.rights.urihttp://creativecommons.org/licenses/by-nc-sa/4.0/en_US
dc.sourceProf. Imperiali via Courtney Crummetten_US
dc.titleDeploying Fluorescent Nucleoside Analogues for High‐Throughput Inhibitor Screeningen_US
dc.typeArticleen_US
dc.identifier.citationSeebald, Leah et al. "Deploying Fluorescent Nucleoside Analogues for High‐Throughput Inhibitor Screening." ChemBioChem 21, 1-2 (January 2020): 108-112 © 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheimen_US
dc.contributor.departmentMassachusetts Institute of Technology. Department of Biologyen_US
dc.contributor.departmentMassachusetts Institute of Technology. Department of Chemistryen_US
dc.relation.journalChemBioChemen_US
dc.eprint.versionAuthor's final manuscripten_US
dc.type.urihttp://purl.org/eprint/type/JournalArticleen_US
eprint.statushttp://purl.org/eprint/status/PeerRevieweden_US
dc.date.updated2020-07-16T17:56:43Z
dspace.date.submission2020-07-16T17:56:46Z
mit.journal.volume21en_US
mit.journal.issue1-2en_US
mit.licenseOPEN_ACCESS_POLICY


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