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Information on EC 3.1.99.B1 - flap endonuclease-1 and Organism(s) Homo sapiens and UniProt Accession P39748
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EC Tree
3.1.99.B1 flap endonuclease-1Specify your search results
Word Map
- 3.1.99.B1
- strand
-
okazaki
-
structure-specific
- pcna
-
exonuclease
- nick
-
single-stranded
-
fork
- nucleases
- duplex
-
5'-flaps
-
long-patch
- endonucleases
-
abasic
- rpa
- ape1
- trinucleotide
-
lp-ber
-
exonucleolytic
-
strand-displacement
-
lagging-strand
-
unannealed
-
pcna-dependent
-
rad9-rad1-hus1
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pcna-binding
-
5'-tails
-
werner
-
3\'-flap
-
incise
-
holliday
- glycosylases
- drug development
The taxonomic range for the selected organisms is: Homo sapiens
The expected taxonomic range for this enzyme is: Archaea, Eukaryota
The expected taxonomic range for this enzyme is: Archaea, Eukaryota
Reaction Schemes
endonucleases that remove 5' DNA sequences from a DNA structure called a DNA flap. The DNA flap structure occurs in double-stranded DNA containing a single-stranded break where the 5' portion of the downstream strand is too long and overlaps the 3' end of the upstream strand. Flap endonucleases cleave the downstream strand of the overlap flap structure precisely after the first base-paired nucleotide, creating a ligatable nick.
endonucleases that remove 5' DNA sequences from a DNA structure called a DNA flap. The DNA flap structure occurs in double-stranded DNA containing a single-stranded break where the 5' portion of the downstream strand is too long and overlaps the 3' end of the upstream strand. Flap endonucleases cleave the downstream strand of the overlap flap structure precisely after the first base-paired nucleotide, creating a ligatable nick
Synonyms
flap endonuclease 1, flap endonuclease, flap endonuclease-1, hfen1, t5fen, fen1 endonuclease, t5 flap endonuclease, affen, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
additional information
additional information
-
the enzyme is a member of the RAD2 family of structure-specific nucleases
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
endonucleases that remove 5' DNA sequences from a DNA structure called a DNA flap. The DNA flap structure occurs in double-stranded DNA containing a single-stranded break where the 5' portion of the downstream strand is too long and overlaps the 3' end of the upstream strand. Flap endonucleases cleave the downstream strand of the overlap flap structure precisely after the first base-paired nucleotide, creating a ligatable nick
in a most likely reaction pathway (very similar to that recently proposed for ribonuclease H, EC 3.1.13.2) the rate-determining step is the SN2-like nucleophilic attack of a water to the scissile phosphate, which occurs concomitantly with its activation by the pro-Rp oxygen of the nucleobase flanking the scissile phosphate. Preactivation of the nucleophilic water. Scheme of different possible reaction mechanisms, overview
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
additional information
?
-
-
FEN1 cleaves substrates bearing telomeric G4 DNA 5'-flaps and is active on substrates bearing telomeric G4 DNA tails, resembling uncapped telomeres, but it is not active on transcribed telomeric G-loops
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-
?
additional information
?
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-
FEN1 contacts the DNA primarily with helix and loop elements. Most interactions are to template strand and terminal flap nts. FEN1 with metal ions has a 1828 A2 interface with product DNA, substrate and product binding structures, detailed overview
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-
?
additional information
?
-
-
Human FEN1 can cleave 5'-flaps bearing structures formed by CTG or CGG repeats, although less efficiently than unstructured flaps. hFEN1 is not affected by the stem-loops formed by CTG repeats interrupting duplex regions adjacent to 5'-flaps, but the enzyme is inhibited by G4 structures formed by CGG repeats in analogous positions. hFEN1 binding causes hypersensitivity near the flap/duplex junction. FEN1 can eliminate structures formed by trinucleotide repeats in the course of replication, relying on endonucleolytic activities
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-
?
additional information
?
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-
substrate recognition and binding, orientation of the flaps into nucleosomes is important for the cleavage, patterns, detailed overview. However, flaps within nucleosomes containing the high-affinity 601 DNA sequence are completely resistant to FEN1 cleavage regardless of orientation. Flaps within such high-affinity nucleosome binding sequences will require ATP-dependent remodeling activity for completion of base excision repair
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-
?
additional information
?
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FEN1 binding to a flap substrate show that the nuclease binds with similar high affinity to the base of a long flap even when the 5'-end is blocked with biotin/streptavidin. However, FEN1 bound to a blocked flap is more sensitive to sequestration by a competing substrate. FEN1 first binds the flap base and then threads the flap through an opening in the protein from the 5'-end to the base for cleavage, substrate interaction mechanism, overview. Short flaps, below two to four nucleotides long, are cleaved without a threading requirement. Direct flap interactions improve FEN1 binding affinity, dissociation rate is slow when FEN1 is bound to a long, unblocked flap
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
additional information
?
-
-
FEN1 cleaves substrates bearing telomeric G4 DNA 5'-flaps and is active on substrates bearing telomeric G4 DNA tails, resembling uncapped telomeres, but it is not active on transcribed telomeric G-loops
-
-
?
additional information
?
-
-
FEN1 contacts the DNA primarily with helix and loop elements. Most interactions are to template strand and terminal flap nts. FEN1 with metal ions has a 1828 A2 interface with product DNA, substrate and product binding structures, detailed overview
-
-
?
additional information
?
-
-
Human FEN1 can cleave 5'-flaps bearing structures formed by CTG or CGG repeats, although less efficiently than unstructured flaps. hFEN1 is not affected by the stem-loops formed by CTG repeats interrupting duplex regions adjacent to 5'-flaps, but the enzyme is inhibited by G4 structures formed by CGG repeats in analogous positions. hFEN1 binding causes hypersensitivity near the flap/duplex junction. FEN1 can eliminate structures formed by trinucleotide repeats in the course of replication, relying on endonucleolytic activities
-
-
?
additional information
?
-
-
substrate recognition and binding, orientation of the flaps into nucleosomes is important for the cleavage, patterns, detailed overview. However, flaps within nucleosomes containing the high-affinity 601 DNA sequence are completely resistant to FEN1 cleavage regardless of orientation. Flaps within such high-affinity nucleosome binding sequences will require ATP-dependent remodeling activity for completion of base excision repair
-
-
?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Mg2+
Sm3+
-
in the active site, the 5'-monophosphate of the cleaved product nt (-1) is coordinated by two Sm3+ ions. Sm1 is coordinated by Asp86, Glu160, and two oxygens of the cleaved 5'-monophosphate. Sm2 is coordinated by Glu160, Asp179, Asp181 and one phosphate oxygen. Asp34, Glu158, and Asp233 interact with Sm3+ via waters
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
-
development of a pair of complementary readout homogeneous assays utilizing fluorogenic donor/quencher and AlphaScreen chemiluminescence strategies for inhibitor screening, overview
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additional information
-
the enzyme is inhibited by G4 structures formed by CGG repeats in duplex regions adjacent to 5'-flaps. hFEN1 5'-flap cleavage is impaired by CTG or CGG repeats
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
-
a telomeric tail, but not a polyA tail, stimulates hFEN1 excision on the opposite strand
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IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0017
NSC-13793
Homo sapiens
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fluorogenic assay and chemoluminescence assay, 22°C, pH not specified in the publication
0.000022
3-hydroxy-5-methyl-1-phenylthieno[2,3-d]pyrimidine-2,4(1H,3H)-dione
Homo sapiens
-
fluorogenic assay, 22°C, pH not specified in the publication
0.000036
3-hydroxy-5-methyl-1-phenylthieno[2,3-d]pyrimidine-2,4(1H,3H)-dione
Homo sapiens
-
chemoluminescence assay, 22°C, pH not specified in the publication
0.00054
aurintricarboxylic acid
Homo sapiens
-
chemoluminescence assay, 22°C, pH not specified in the publication
0.00059
aurintricarboxylic acid
Homo sapiens
-
pH and temperature not specified in the publication
0.00063
aurintricarboxylic acid
Homo sapiens
-
fluorogenic assay, 22°C, pH not specified in the publication
0.13
NSC-13744
Homo sapiens
-
chemoluminescence assay, 22°C, pH not specified in the publication
0.00066
NSC-13755
Homo sapiens
-
chemoluminescence assay, 22°C, pH not specified in the publication
0.012
NSC-15596
Homo sapiens
-
chemoluminescence assay, 22°C, pH not specified in the publication
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
additional information
FEN1 is overexpressed in all proliferative tissues
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
physiological function
Flap endonucleases (FENs) are nucleic acid hydrolyzing enzymes in charge of excising 5'-small DNA and RNA fragments (flaps) protruding from nucleic acid structures during the lagging strand DNA replication or the longpatch base excision repair (LP-BER) processes. Important role of FENs in maintaining nucleic acid fidelity and cell proliferation, high levels of FEN1 are believed to support cancer cell hyperproliferation
additional information
mixed quantum-classical (QM/MM) metadynamics and umbrella sampling free energy calculations are employed on a reconstructed reactive hFEN/double strand (ds) DNA adduct for an atomistic and energetic rendering of the enzymatic catalysis promoted by the human FEN1, structure-function analysis, overview. The enzymatic phosphate hydrolysis proceeds as an SN2-like nucleophilic attack on the scissile phosphate performed by an hydroxide ion, which is typically formed upon water activation
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
purified recombinant His-tagged FEN1 mutant DELTA 336 and FEN1 mutant DELTA 336 D181A, X-ray diffraction structure determination and analysis
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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
D181A
-
construction of a FEN1 mutant DELTA 336 truncated after residue 336 that bears an additional point mutation at residue 181. FEN1 DELTA336 removes only the flexible, protruding PCNA binding motif and encompasses the entire catalytic domain, and the active site D181A mutation severely retards incision. The DNA contains competing base pairing purine-pyrimidine pairs at the DNA junction, but the C-A mismatch favors 1 nt 3' flap formation
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant His6-tagged FEN1 mutant DELTA 336 and FEN1 mutant DELTA 336 D181A from Escherichia coli
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CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expression of His6-tagged FEN1 mutant DELTA 336 and FEN1 mutant DELTA 336 D181A in Escherichia coli
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
drug development
-
inhibitors of the enzyme carry a potential as enhancers of DNA-interactive anticancer drugs
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Vallur, A.C.; Maizels, N.
Complementary roles for exonuclease 1 and Flap endonuclease 1 in maintenance of triplet repeats
J. Biol. Chem.
285
28514-28519
2010
Homo sapiens
Vallur, A.C.; Maizels, N.
Distinct activities of exonuclease 1 and flap endonuclease 1 at telomeric g4 DNA
PLoS ONE
5
e8908
2010
Homo sapiens
Tsutakawa, S.E.; Classen, S.; Chapados, B.R.; Arvai, A.S.; Finger, L.D.; Guenther, G.; Tomlinson, C.G.; Thompson, P.; Sarker, A.H.; Shen, B.; Cooper, P.K.; Grasby, J.A.; Tainer, J.A.
Human flap endonuclease structures, DNA double-base flipping, and a unified understanding of the FEN1 superfamily
Cell
145
198-211
2011
Homo sapiens
Gloor, J.W.; Balakrishnan, L.; Bambara, R.A.
Flap endonuclease 1 mechanism analysis indicates flap base binding prior to threading
J. Biol. Chem.
285
34922-34931
2010
Homo sapiens
Jagannathan, I.; Pepenella, S.; Hayes, J.J.
Activity of FEN1 endonuclease on nucleosome substrates is dependent upon DNA sequence but not flap orientation
J. Biol. Chem.
286
17521-17529
2011
Homo sapiens
Fortini, B.K.; Pokharel, S.; Polaczek, P.; Balakrishnan, L.; Bambara, R.A.; Campbell, J.L.
Characterization of the endonuclease and ATP-dependent flap endo/exonuclease of Dna2
J. Biol. Chem.
286
23763-23770
2011
Homo sapiens
Dorjsuren, D.; Kim, D.; Maloney, D.J.; Wilson, D.M.; Simeonov, A.
Complementary non-radioactive assays for investigation of human flap endonuclease 1 activity
Nucleic Acids Res.
39
e11
2011
Homo sapiens
Sgrignani, J.; Magistrato, A.
QM/MM MD simulations on the enzymatic pathway of the human flap endonuclease (hFEN1) elucidating common cleavage pathways to RNase H enzymes
ACS Catal.
5
3864-3875
2015
Homo sapiens (P39748)
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