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dc.contributor.authorCraggs, Timothy David
dc.contributor.authorBrenlla, Alfonso
dc.contributor.authorHutton, Richard David
dc.contributor.authorWhite, Malcolm F
dc.contributor.authorPenedo, Carlos
dc.identifier.citationCraggs , T D , Brenlla , A , Hutton , R D , White , M F & Penedo , C 2014 , ' Single-molecule characterization of Fen1 and Fen1/PCNA complexes acting on flap substrates ' , Nucleic Acids Research , vol. 42 , no. 3 , pp. 1857-1872 .
dc.identifier.otherORCID: /0000-0003-1543-9342/work/47136108
dc.identifier.otherORCID: /0000-0002-5807-5385/work/74872759
dc.descriptionThis article was made OA through RCUK block grant funds.en
dc.description.abstractFlap endonuclease 1 (Fen1) is a highly conserved structure-specific nuclease that catalyses a specific incision to remove 5′ flaps in double-stranded DNA substrates. Fen1 plays an essential role in key cellular processes, such as DNA replication and repair, and mutations that compromise Fen1 expression levels or activity have severe health implications in humans. The nuclease activity of Fen1 and other FEN family members can be stimulated by processivity clamps such as proliferating cell nuclear antigen (PCNA); however, the exact mechanism of PCNA activation is currently unknown. Here, we have used a combination of ensemble and single-molecule Förster resonance energy transfer together with protein-induced fluorescence enhancement to uncouple and investigate the substrate recognition and catalytic steps of Fen1 and Fen1/PCNA complexes. We propose a model in which upon Fen1 binding, a highly dynamic substrate is bent and locked into an open flap conformation where specific Fen1/DNA interactions can be established. PCNA enhances Fen1 recognition of the DNA substrate by further promoting the open flap conformation in a step that may involve facilitated threading of the 5′ ssDNA flap. Merging our data with existing crystallographic and molecular dynamics simulations we provide a solution-based model for the Fen1/PCNA/DNA ternary complex.
dc.relation.ispartofNucleic Acids Researchen
dc.subjectFlap endonuclease 1 (Fen1)en
dc.subjectProliferating cell nuclear antigen (PCNA)en
dc.subjectFörster resonance energy transferen
dc.subjectFluorescence enhancementen
dc.subjectQH426 Geneticsen
dc.titleSingle-molecule characterization of Fen1 and Fen1/PCNA complexes acting on flap substratesen
dc.typeJournal articleen
dc.contributor.institutionUniversity of St Andrews. School of Physics and Astronomyen
dc.contributor.institutionUniversity of St Andrews. Biomedical Sciences Research Complexen
dc.contributor.institutionUniversity of St Andrews. School of Biologyen
dc.description.statusPeer revieweden

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