title: Sequential and multistep substrate interrogation provides the scaffold for specificity in human flap endonuclease 1.
abstract: Human flap endonuclease 1 (FEN1), one of the structure-specific 5' nucleases, is integral in replication, repair, and recombination of cellular DNA. The 5' nucleases share significant unifying features yet cleave diverse substrates at similar positions relative to 5' end junctions. Using single-molecule Förster resonance energy transfer, we find a multistep mechanism that verifies all substrate features before inducing the intermediary-DNA bending step that is believed to unify 5' nuclease mechanisms. This is achieved by coordinating threading of the 5' flap of a nick junction into the conserved capped-helical gateway, overseeing the active site, and bending by binding at the base of the junction. We propose that this sequential and multistep substrate recognition process allows different 5' nucleases to recognize different substrates and restrict the induction of DNA bending to the last common step. Such mechanisms would also ensure the protection of DNA junctions from nonspecific bending and cleavage.
authors.affiliation: Division of Biological and Environmental Sciences and Engineering, King Abdullah University of Science and Technology, Thuwal 23955, Saudi Arabia.
authors.forename: Mohamed A
authors.initials: MA
authors.lastname: Sobhy
authors.forename: Luay I
authors.initials: LI
authors.lastname: Joudeh
authors.forename: Xiaojuan
authors.initials: X
authors.lastname: Huang
authors.forename: Masateru
authors.initials: M
authors.lastname: Takahashi
authors.forename: Samir M
authors.initials: SM
authors.lastname: Hamdan
mesh_terms: D004247:DNA / Q000737:chemistry / Q000378:metabolism; D004261:DNA Replication; D045585:Flap Endonucleases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D013379:Substrate Specificity
chemical_list: D004247:DNA; D045585:Flap Endonucleases; C468317:FEN1 protein, human