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dc.contributor.authorKrivoy, Andrey
dc.contributor.authorRutkauskas, Marius
dc.contributor.authorKuznedelov, Konstantin
dc.contributor.authorMusharova, Olga
dc.contributor.authorRouillon, Christophe
dc.contributor.authorSeverinov, Konstantin
dc.contributor.authorSeidel, Ralf
dc.date.accessioned2018-04-20T11:30:11Z
dc.date.available2018-04-20T11:30:11Z
dc.date.issued2018-05-04
dc.identifier.citationKrivoy , A , Rutkauskas , M , Kuznedelov , K , Musharova , O , Rouillon , C , Severinov , K & Seidel , R 2018 , ' Primed CRISPR adaptation in Escherichia coli cells does not depend on conformational changes in the Cascade effector complex detected in vitro ' , Nucleic Acids Research , vol. 46 , no. 8 , pp. 4087-4098 . https://doi.org/10.1093/nar/gky219en
dc.identifier.issn0305-1048
dc.identifier.otherPURE: 252882351
dc.identifier.otherPURE UUID: 70e5054d-8d48-4a60-8e57-0f767ab72e0d
dc.identifier.otherPubMed: 29596641
dc.identifier.otherScopus: 85052592154
dc.identifier.otherWOS: 000431895800031
dc.identifier.urihttp://hdl.handle.net/10023/13172
dc.descriptionSkoltech Ph.D. program in the Life Sciences (to A.K.); European Research Council consolidator grant [GA 724863 to R.S].; NIH [R01 GM10407] and Russian Science Foundation [14-14-00988] grants to K.S.; UMNIK grant 8115GU/2015 to O.M., and institutional support from Skoltech to K.S. Funding for open access charge: Skolkovo Institute of Science and Technology internal funding.en
dc.description.abstractIn type I CRISPR-Cas systems, primed adaptation of new spacers into CRISPR arrays occurs when the effector Cascade-crRNA complex recognizes imperfectly matched targets that are not subject to efficient CRISPR interference. Thus, primed adaptation allows cells to acquire additional protection against mobile genetic elements that managed to escape interference. Biochemical and biophysical studies suggested that Cascade-crRNA complexes formed on fully matching targets (subject to efficient interference) and on partially mismatched targets that promote primed adaption are structurally different. Here, we probed Escherichia coli Cascade-crRNA complexes bound to matched and mismatched DNA targets using a magnetic tweezers assay. Significant differences in complex stabilities were observed consistent with the presence of at least two distinct conformations. Surprisingly, in vivo analysis demonstrated that all mismatched targets stimulated robust primed adaptation irrespective of conformational states observed in vitro. Our results suggest that primed adaptation is a direct consequence of a reduced interference efficiency and/or rate and is not a consequence of distinct effector complex conformations on target DNA.
dc.format.extent12
dc.language.isoeng
dc.relation.ispartofNucleic Acids Researchen
dc.rights© The Author(s) 2018. Published by Oxford University Press on behalf of Nucleic Acids Research. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.comen
dc.subjectQH301 Biologyen
dc.subjectNDASen
dc.subject.lccQH301en
dc.titlePrimed CRISPR adaptation in Escherichia coli cells does not depend on conformational changes in the Cascade effector complex detected in vitroen
dc.typeJournal articleen
dc.description.versionPublisher PDFen
dc.contributor.institutionUniversity of St Andrews.School of Biologyen
dc.identifier.doihttps://doi.org/10.1093/nar/gky219
dc.description.statusPeer revieweden


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