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Information on EC 3.6.4.12 - DNA helicase and Organism(s) Homo sapiens and UniProt Accession Q9BX63

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IUBMB Comments
DNA helicases utilize the energy from ATP hydrolysis to unwind double-stranded DNA. Some of them unwind duplex DNA with a 3' to 5' polarity [1,3,5,8], others show 5' to 3' polarity [10,11,12,13] or unwind DNA in both directions [14,15]. Some helicases unwind DNA as well as RNA [9,10]. May be identical with EC 3.6.4.13 (RNA helicase).
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This record set is specific for:
Homo sapiens
UNIPROT: Q9BX63
Word Map
The taxonomic range for the selected organisms is: Homo sapiens
The enzyme appears in selected viruses and cellular organisms
Reaction Schemes
Synonyms
recq helicase, twinkle, fancj, dnab helicase, recq1, rtel1, mcm complex, wrn helicase, minichromosome maintenance protein, helicase ii, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
ATP-dependent DNA helicase Q1
285427
-
BACH1 helicase
285412
-
BLM protein
247
-
Bloom syndrome protein
304137
-
BRCA1-associated C-terminal helicase
285412
-
C10orf2
285414
gene name
ChlR1 helicase
304139
-
DDX25
247
-
DNA helicase Rtel1
247
-
DNA helicase VI
247
-
FANCJ
298130
-
Fanconi Anemia Group J Protein
298130
-
GRTH/DDX25
247
-
HDH IV
247
-
HDHB
247
-
hPif1
285426
-
human DNA helicase B
247
-
Mcm4/6/7 complex
247
-
PIF1
285426
-
PIF1 helicase
285426
-
RecQ1
285427
-
Rtel1
247
-
SF2 helicase
285427, 304137
-
telomere length regulator
247
-
TWINKLE
TWINKLE DNA helicase
247
-
Werner Syndrome helicase
285411
-
Werner syndrome RecQ helicase
285411
-
WRN
285411
-
WRN helicase
285411
-
WRN protein
285411
-
WRN RecQ helicase
285411
-
XPD
247
-
SYSTEMATIC NAME
IUBMB Comments
ATP phosphohydrolase (DNA helix unwinding)
DNA helicases utilize the energy from ATP hydrolysis to unwind double-stranded DNA. Some of them unwind duplex DNA with a 3' to 5' polarity [1,3,5,8], others show 5' to 3' polarity [10,11,12,13] or unwind DNA in both directions [14,15]. Some helicases unwind DNA as well as RNA [9,10]. May be identical with EC 3.6.4.13 (RNA helicase).
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ATP + H2O
ADP + phosphate
show the reaction diagram
-
-
-
?
ATP + H2O
ADP + phosphate
show the reaction diagram
dATP + H2O
dADP + phosphate
show the reaction diagram
-
ATP-dependent DNA unwinding enzyme. HDH VI unwinds exclusively DNA duplexes with an annealed portion smaller than32 bp and prefers a replication fork-like structure of the substrate. It cannot unwind blunt-end duplexes and is inactive also on DNA-RNA or RNA-RNA hybrids. HDH VI unwinds DNA unidirectionally by moving in the 3' to 5' direction along the bound strand. ATP and dATP are equally good substrates
-
-
?
dTTP + H2O
dTDP + phosphate
show the reaction diagram
TWINKLE is a DNA helicase with 5' to 3' directionality. The enzyme needs a stretch of 10 nucleotides of single-stranded DNA on the 5'-side of the duplex to unwind duplex DNA. In addition, helicase activity is not observed unless a short single-stranded 3'-tail is present. UTP efficiently supports DNA unwinding. ATP, GTP, and dTTP are less effective
-
-
?
GTP + H2O
GDP + phosphate
show the reaction diagram
TWINKLE is a DNA helicase with 5' to 3' directionality. The enzyme needs a stretch of 10 nucleotides of single-stranded DNA on the 5'-side of the duplex to unwind duplex DNA. In addition, helicase activity is not observed unless a short single-stranded 3'-tail is present. UTP efficiently supports DNA unwinding. ATP, GTP, and dTTP are less effective
-
-
?
UTP + H2O
UDP + phosphate
show the reaction diagram
additional information
?
-
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
ATP + H2O
ADP + phosphate
show the reaction diagram
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Fe-S
-
-
H2O2
-
the iron-sulfur cluster in the N-terminus of the enzyme, RtelN, residues 1-312, is sensitive to hydrogen peroxide and nitric oxide, indicating that reactive oxygen/nitrogen species may modulate the DNA helicase activity of Rtel1 via modification of its iron-sulfur cluster but not significantly affect the DNA binding activity of RtelN
KCl
-
optimal concentration: 100 mM. Inhibition at 200 mM
MgUTP
-
MgUTP stabilizes hexamers over higher oligomers
nitric oxide
-
the iron-sulfur cluster in the N-terminus of the enzyme, RtelN, residues 1-312, is sensitive to hydrogen peroxide and nitric oxide, indicating that reactive oxygen/nitrogen species may modulate the DNA helicase activity of Rtel1 via modification of its iron-sulfur cluster but not significantly affect the DNA binding activity of RtelN
additional information
-
the N-terminal domain of DNA helicase Rtel1, residues 1-312, contains a redox active iron-sulfur cluster
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
ATPgammaS
-
-
benzoquinoquinoxaline
BQQ, inhibits ChlR1 triplex DNA unwinding activity
EDTA
-
5 mM, complete inhibition
KCl
-
optimal concentration: 100 mM. Inhibition at 200 mM
Mg2+
-
required, optimal concentration: 0.8 mM. Inhibition at 4 mM
replication protein A
-
inhibits unwinding and annealing activities
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streptavidin
-
the enzyme is completely blocked by streptavidin bound to the 3'-ssDNA tail 6 nucleotides upstream of the single-stranded/double-stranded DNA junction. The enzyme efficiently unwinds the forked duplex with streptavidin bound just upstream of the junction, suggesting that the enzyme recognizes elements of the fork structure to initiate unwinding
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Tim protein
-
the Tim protein alone also decreases the ATPase and DNA helicase activities of the Mcm 4/6/7 complex
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Tim-Tipin complex
-
the addition of the human Tim-Tipin complex inhibits both ATPase and DNA helicase activities of the Mcm4/6/7 complex. The ATPase activity in the presence of ssDNA appears to be affected more severely compared with that seen in the absence of ssDNA. The Tim protein alone also decreases the ATPase and DNA helicase activities of the Mcm 4/6/7 complex, whereas Tipin does not
-
additional information
enzyme ChlR1 fails to unwind the triplex substrate in the absence of ATP or in the presence of ADP or ATPgammaS
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Double-stranded DNA
-
the enzyme is strongly stimulated by either single- or double-stranded DNA
mitochondrial single-stranded DNA-binding protein
-
stimultes the enzyme
-
replication protein A
-
single-stranded DNA
additional information
no significant stimulation by Escherichia coli ssDNA-binding protein
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.89 - 1.9
ATP
0.0038 - 4.55
ATP
1.4 - 3.4
UTP
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.07 - 43.3
ATP
0.66 - 11.25
ATP
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.54 - 1.83
ATP
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.4
assay at
7.8
-
assay at
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30
assay at
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
UniProt
Manually annotated by BRENDA team
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
-
GRTH is a negative regulator of apoptosis in spermatocytes and promotes the progress of spermatogenesis
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
malfunction
-
HDHB knockdown impairs recovery from replication stress
physiological function
additional information
structure comparisons of SF1 and SF2 helicases, SF1 and SF2 helicase domains structures and substrate-bound SF1 and SF2 helicase structures, structure-function relationship, overview
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
Sequence
FANCJ_HUMAN
1249
0
140867
Swiss-Prot
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
70000
-
x * 70000, SDS-PAGE
71000
-
1 * 71000, SDS-PAGE
72000
-
SDS-PAGE
116000
6 * 116000, SDS-PAGE
128000
-
gel filtration
180000
-
x * 180000, SDS-PAGE
600000
gel filtration
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
FANCJ exists as molecular weight species corresponding to a monomer and a dimer, and the dimeric form displays a higher specific activity for ATPase and helicase, as well as greater DNA binding
monomer
FANCJ exists as molecular weight species corresponding to a monomer and a dimer, and the dimeric form displays a higher specific activity for ATPase and helicase, as well as greater DNA binding
hexamer
monomer
oligomer
-
recombinant TWINKLE assembles into hexamers and higher oligomers, and addition of MgUTP stabilizes hexamers over higher oligomers
additional information
structure-function relationship
CRYSTALLIZATION/commentary
ORGANISM
UNIPROT
LITERATURE
crystal structure analysis of the helicase domain from the SF2A DNA helicase BLM bound to partial-duplex DNA, PDB ID 4O3M, and structure PDB ID 4CGZ
crystal structure analysis, PDB ID 2WWY
purified RECQ1T1 protein is crystallized in the presence of ATP-gammaS and oligonucleotides by vapor diffusion from sitting drops equilibrated against 0.2 M sodium bromide, 20% PEG 3350, 10% ethylene glycol, 0.1 M bis-Tris propane (pH 7.5). Crystal structure of a truncated form (RECQ1(T1)) of the human RECQ1 protein with MgADP2-
-
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
A349P
expression of a FANCJ mutant protein with a pathogenic A349P amino acid substitution in a wild-type background exerts a dominant negative effect on resistance to DNA cross-linking agents or TMS
DELTAQ25
mutant bearing a deletion of Q25 shows a similar phenotype as Q25A
K52R
mutant is seriously compromised in its ATPase activity, kcat (ATP) highly decreased compared to wild-type
Q25A
Q25A mutation of the invariant glutamine in the Q motif abolishes its ability to complement cisplatin or telomestatin sensitivity of a fancj null cell line and exerts a dominant negative effect. Mutant shows impaired FANCJ helicase activity and ATPase activity but displays ATP binding and temperature-induced unfolding transition similar to wild-type. Mutant exists only as a monomer. Km (ATP) increased compared to wild-type, kcat 10fold decreased compared to wild-type
A318T
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) comparable to wild-type
A359T
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) decreased compared to wild-type, mutant shows a 3-4fold increase in Kd (dsDNA) value compared to wild-type, mutant is much more sensitive to heat inactivation than wild-type
A475P
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) decreased compared to wild-type, mutant shows lowest kcat (ATP) value
F485L
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) decreased compared to wild-type, mutant shows a 3-4fold increase in Kd (dsDNA) value compared to wild-type, mutant is much more sensitive to heat inactivation than wild-type
I367T
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) slightly decreased compared to wild-type, mutant shows a higher Km (ATP) and kcat value compared to wild-type
K319E
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) comparable to wild-type
K319T
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) decreased compared to wild-type
K421A
-
the Walker A mutant K421A of TWINKLE fails to unwind dsDNA
K50R
site-directed mutagenesis, ATPase dead mutant, fails completely to unwind triplex substrates
K897del
site-directed mutagenesis, the mutant fails in unwinding the DNA substrates
L381P
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) increased compared to wild-type, mutant shows a higher Km (ATP) and kcat value compared to wild-type
P335L
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) decreased compared to wild-type
R112H
-
most common mutations in TTD patients, amino acid substitution R112H, is localized in the Fe-S domain of XPD just before the first conserved cysteine residue. Missense mutation results in a complete loss of XPD helicase activity and a reduced basal transcription activity of the TFIIH complex
R303W
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) comparable to wild-type, mutant is much more sensitive to heat inactivation than wild-type
R334Q
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) decreased compared to wild-type
R354P
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) decreased compared to wild-type
R357P
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) slightly decreased compared to wild-type
R374Q
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) decreased compared to wild-type, mutant shows a higher Km (ATP) value compared to wild-type, mutant shows a 3-4fold increase in Kd (dsDNA) and a 2fold increase in Kd (ssDNA) value compared to wild-type, mutant is much more sensitive to heat inactivation than wild-type
S369P
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) decreased compared to wild-type
S369Y
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) increased compared to wild-type, mutant shows a 3-4fold increase in Kd (dsDNA) value compared to wild-type, mutant is much more sensitive to heat inactivation than wild-type
T457I
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) decreased compared to wild-type
W315L
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) slightly decreased compared to wild-type, mutant is much more sensitive to heat inactivation than wild-type
W474C
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) decreased compared to wild-type
Y508C
-
mutation associated with mitochondrial disease does not cause profound defects in DNA binding, DNA helicase function, or ATPase activity, kcat/Km (ATP) decreased compared to wild-type, mutant shows a 3-4fold increase in Kd (dsDNA) value compared to wild-type
additional information
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
42
-
wild-type protein has a half-liife if 7.9 min at 42°C
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
enzyme activity is destroyed if trypsin is included in the reaction
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
4°C, DNA helicase VI loses 90% of its activity in 24 h
-
PURIFICATION/commentary
ORGANISM
UNIPROT
LITERATURE
a construct (RECQ1(T1)) encompassing amino acids 49–616 (of 649) of RECQ1, followed by a C-terminal tag of 22 aa is produced in Escherichia coli and purified to more than 95% homogeneity
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full-length PIF1 with a 6* histidine tag at the N-terminus, a C-terminal truncated form (PIF1N) and a N-terminal truncated form (PIF1C)
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generation of recombinant baculovirus encoding the human TWINKLE gene, expression in insect cells
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recombinant
-
streamlined purification for the production of near-homogeneous and high yield recombinant forms of the human mitochondrial DNA helicase, minimizing the number of steps and the time elapsed for purification
-
using Ni-NTA chromatography
-
CLONED/commentary
ORGANISM
UNIPROT
LITERATURE
expressed in insect cells using the baculovirus expression system
a construct (RECQ1(T1)) encompassing amino acids 49–616 (of 649) of RECQ1, followed by a C-terminal tag of 22 aa is produced in Escherichia coli
-
expressed in Hi5 insect cells
-
expressed in SF9 cells as His-tagged fusion proteins lacking the N-terminal mitochondrial targeting signal using the baculovirus expression system
-
expression in Escherichia coli
human hPif1 (nuclear form amino acids 1–641) and the hPif helicase domain (hPifHD, amino acid residues 206–620) are cloned as a fusion protein with glutathione S-transferase in pET11c. GST-hPifHD is expressed in Escherichia coli BL21(DE3) cells
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overexpressed in Escherichia coli as His-tagged fusion proteins
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overexpression of an oligohistidine-tagged version of the BLM gene product in Saccharomyces cerevisiae
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recombinant expression of FLAG3-tagged wild-type and mutant enzymes in HEK293T cells
recombinant expression of the N-terminal domain of Rtel1, RtelN, residues 1-312, in Escherichia coli strain BL21
-
REF.
AUTHORS