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dc.contributor.authorDowling, Daniel P.
dc.contributor.authorKung, Yan
dc.contributor.authorCroft, Anna K.
dc.contributor.authorTaghizadeh, Koli
dc.contributor.authorKelly, Wendy L.
dc.contributor.authorWalsh, Christopher T.
dc.contributor.authorDrennan, Catherine L.
dc.date.accessioned2017-05-12T14:47:00Z
dc.date.available2017-05-12T14:47:00Z
dc.date.issued2016-10
dc.date.submitted2016-05
dc.identifier.issn0027-8424
dc.identifier.issn1091-6490
dc.identifier.urihttp://hdl.handle.net/1721.1/109039
dc.description.abstractEpothilones are thiazole-containing natural products with anticancer activity that are biosynthesized by polyketide synthase (PKS)-nonribosomal peptide synthetase (NRPS) enzymes EpoA–F. A cyclization domain of EpoB (Cy) assembles the thiazole functionality from an acetyl group and l-cysteine via condensation, cyclization, and dehydration. The PKS carrier protein of EpoA contributes the acetyl moiety, guided by a docking domain, whereas an NRPS EpoB carrier protein contributes l-cysteine. To visualize the structure of a cyclization domain with an accompanying docking domain, we solved a 2.03-Å resolution structure of this bidomain EpoB unit, comprising residues M1-Q497 (62 kDa) of the 160-kDa EpoB protein. We find that the N-terminal docking domain is connected to the V-shaped Cy domain by a 20-residue linker but otherwise makes no contacts to Cy. Molecular dynamic simulations and additional crystal structures reveal a high degree of flexibility for this docking domain, emphasizing the modular nature of the components of PKS-NRPS hybrid systems. These structures further reveal two 20-Å-long channels that run from distant sites on the Cy domain to the active site at the core of the enzyme, allowing two carrier proteins to dock with Cy and deliver their substrates simultaneously. Through mutagenesis and activity assays, catalytic residues N335 and D449 have been identified. Surprisingly, these residues do not map to the location of the conserved HHxxxDG motif in the structurally homologous NRPS condensation (C) domain. Thus, although both C and Cy domains have the same basic fold, their active sites appear distinct.en_US
dc.description.sponsorshipNational Institute of Environmental Health Sciences (P30-ES002109)en_US
dc.language.isoen_US
dc.publisherNational Academy of Sciences (U.S.)en_US
dc.relation.isversionofhttp://dx.doi.org/10.1073/pnas.1608615113en_US
dc.rightsArticle is made available in accordance with the publisher's policy and may be subject to US copyright law. Please refer to the publisher's site for terms of use.en_US
dc.sourcePNASen_US
dc.titleStructural elements of an NRPS cyclization domain and its intermodule docking domainen_US
dc.typeArticleen_US
dc.identifier.citationDowling, Daniel P.; Kung, Yan; Croft, Anna K.; Taghizadeh, Koli; Kelly, Wendy L.; Walsh, Christopher T. and Drennan, Catherine L. “Structural Elements of an NRPS Cyclization Domain and Its Intermodule Docking Domain.” Proceedings of the National Academy of Sciences 113, no. 44 (October 2016): 12432–12437. © 2016 National Academy of Sciencesen_US
dc.contributor.departmentMassachusetts Institute of Technology. Department of Biologyen_US
dc.contributor.departmentMassachusetts Institute of Technology. Department of Chemistryen_US
dc.contributor.mitauthorDowling, Daniel P.
dc.contributor.mitauthorKung, Yan
dc.contributor.mitauthorDrennan, Catherine L.
dc.relation.journalProceedings of the National Academy of Sciencesen_US
dc.eprint.versionFinal published versionen_US
dc.type.urihttp://purl.org/eprint/type/JournalArticleen_US
eprint.statushttp://purl.org/eprint/status/PeerRevieweden_US
dspace.orderedauthorsDowling, Daniel P.; Kung, Yan; Croft, Anna K.; Taghizadeh, Koli; Kelly, Wendy L.; Walsh, Christopher T.; Drennan, Catherine L.en_US
dspace.embargo.termsNen_US
dc.identifier.orcidhttps://orcid.org/0000-0001-5486-2755
dspace.mitauthor.errortrue
mit.licensePUBLISHER_POLICYen_US


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