title: Human flap endonuclease structures, DNA double-base flipping, and a unified understanding of the FEN1 superfamily.
abstract: Flap endonuclease (FEN1), essential for DNA replication and repair, removes RNA and DNA 5' flaps. FEN1 5' nuclease superfamily members acting in nucleotide excision repair (XPG), mismatch repair (EXO1), and homologous recombination (GEN1) paradoxically incise structurally distinct bubbles, ends, or Holliday junctions, respectively. Here, structural and functional analyses of human FEN1:DNA complexes show structure-specific, sequence-independent recognition for nicked dsDNA bent 100° with unpaired 3' and 5' flaps. Above the active site, a helical cap over a gateway formed by two helices enforces ssDNA threading and specificity for free 5' ends. Crystallographic analyses of product and substrate complexes reveal that dsDNA binding and bending, the ssDNA gateway, and double-base unpairing flanking the scissile phosphate control precise flap incision by the two-metal-ion active site. Superfamily conserved motifs bind and open dsDNA; direct the target region into the helical gateway, permitting only nonbase-paired oligonucleotides active site access; and support a unified understanding of superfamily substrate specificity.
authors.affiliation: Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA.
authors.forename: Susan E
authors.initials: SE
authors.lastname: Tsutakawa
authors.forename: Scott
authors.initials: S
authors.lastname: Classen
authors.forename: Brian R
authors.initials: BR
authors.lastname: Chapados
authors.forename: Andrew S
authors.initials: AS
authors.lastname: Arvai
authors.forename: L David
authors.initials: LD
authors.lastname: Finger
authors.forename: Grant
authors.initials: G
authors.lastname: Guenther
authors.forename: Christopher G
authors.initials: CG
authors.lastname: Tomlinson
authors.forename: Peter
authors.initials: P
authors.lastname: Thompson
authors.forename: Altaf H
authors.initials: AH
authors.lastname: Sarker
authors.forename: Binghui
authors.initials: B
authors.lastname: Shen
authors.forename: Priscilla K
authors.initials: PK
authors.lastname: Cooper
authors.forename: Jane A
authors.initials: JA
authors.lastname: Grasby
authors.forename: John A
authors.initials: JA
authors.lastname: Tainer
mesh_terms: D000595:Amino Acid Sequence; D020134:Catalytic Domain; D004247:DNA / Q000378:metabolism; D004252:DNA Mutational Analysis; D005090:Exodeoxyribonucleases / Q000737:chemistry / Q000378:metabolism; D045585:Flap Endonucleases / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016415:Sequence Alignment; D013379:Substrate Specificity
chemical_list: D004247:DNA; D005090:Exodeoxyribonucleases; D045585:Flap Endonucleases; C468317:FEN1 protein, human