Information on EC 2.7.7.7 - DNA-directed DNA polymerase and Organism(s) Homo sapiens and UniProt Accession Q9Y253

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UNIPROT: Q9Y253
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The taxonomic range for the selected organisms is: Homo sapiens

The enzyme appears in selected viruses and cellular organisms

EC NUMBER
COMMENTARY hide
2.7.7.7
-
RECOMMENDED NAME
GeneOntology No.
DNA-directed DNA polymerase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
a 2'-deoxyribonucleoside 5'-triphosphate + DNAn = diphosphate + DNAn+1
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
nucleotidyl group transfer
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Metabolic pathways
-
-
Purine metabolism
-
-
Pyrimidine metabolism
-
-
SYSTEMATIC NAME
IUBMB Comments
deoxynucleoside-triphosphate:DNA deoxynucleotidyltransferase (DNA-directed)
Catalyses DNA-template-directed extension of the 3'- end of a DNA strand by one nucleotide at a time. Cannot initiate a chain de novo. Requires a primer, which may be DNA or RNA. See also EC 2.7.7.49 RNA-directed DNA polymerase.
CAS REGISTRY NUMBER
COMMENTARY hide
9012-90-2
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
SwissProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
DNA polymerase eta-deficient cells show strong activation of downstream DNA damage responses including ataxia-telangiectasia mutated and Rad3-related protein signaling and accumulate strand breaks as result of replication fork collapse
physiological function
DNA polymerase eta is a key protein in translesion synthesis in human cells, it is a low-fidelity enzyme when copying undamaged DNA but can carry out error-free translesion synthesis at sites of UV-induced dithymine cyclobutane pyrimidine dimmers. DNA polymerase eta plays an important role in preventing genome instability after UV- and cisplatin-induced DNA damage
evolution
malfunction
metabolism
physiological function
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
deoxynucleoside triphosphate + DNAn
diphosphate + DNAn+1
show the reaction diagram
-
-
-
?
2-hydroxy-2'-deoxyadenosine 5'-triphosphate + DNAn
diphosphate + DNAn+1
show the reaction diagram
-
DNA polymerase eta incorporates 2-hydroxy-2'-deoxyadenosine 5'-triphosphate opposite template G during DNA synthesis
-
-
?
8-hydroxy-2'-deoxyguanosine 5'-triphosphate + DNAn
diphosphate + DNAn+1
show the reaction diagram
-
DNA polymerase eta incorporates 8-hydroxy-2'-deoxyguanosine 5'-triphosphate opposite template A and slightly opposite template C during DNA synthesis
-
-
?
dATP + DNAn
diphosphate + DNAn+1
show the reaction diagram
-
with activated calf thymus DNA
-
-
?
dCTP + DNAn
diphosphate + DNAn+1
show the reaction diagram
deoxynucleoside triphosphate + DNAn
diphosphate + DNAn+1
show the reaction diagram
dGTP + DNAn
diphosphate + DNAn+1
show the reaction diagram
-
with activated calf thymus DNA
-
-
?
dTTP + DNAn
diphosphate + DNAn+1
show the reaction diagram
additional information
?
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
deoxynucleoside triphosphate + DNAn
diphosphate + DNAn+1
show the reaction diagram
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Na+
-
pols beta, iota and zeta exhibit maximal activity under conditions of moderate NaCl concentrations of 20-60 mM
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
(2E)-2-(3-nitro-4-[[6-(trifluoromethyl)pyridin-3-yl]sulfanyl]benzylidene)-5-thioxo-1,3-thiazolidin-4-one
-
-
(2E)-2-(4-chloro-3-ethylbenzylidene)-5-thioxo-1,3-thiazolidin-4-one
-
-
(2E)-2-(pentafluorobenzylidene)-5-thioxodihydrothiophen-3(2H)-one
-
-
(2E)-2-[4-(2-hydroxyethyl)-3-nitrobenzylidene]-5-thioxo-1,3-thiazolidin-4-one
-
-
(5Z)-5-(4-chloro-3-nitrobenzylidene)-2-thioxo-1,3-thiazolidin-4-one
-
-
(5Z)-5-[3-bromo-4-[(4-fluorobenzyl)oxy]benzylidene]-1,3-thiazolidine-2,4-dione
-
-
(5Z)-5-[3-bromo-4-[(4-fluorophenyl)sulfanyl]benzylidene]-2-thioxo-1,3-thiazolidin-4-one
-
-
(5Z)-5-[3-nitro-4-(pyridin-3-ylsulfanyl)benzylidene]-2-thioxo-1,3-thiazolidin-4-one
-
-
(5Z)-5-[4-(4-methylphenoxy)-3-nitrobenzylidene]-2-thioxo-1,3-thiazolidin-4-one
-
-
(5Z)-5-[4-(cyclohexylsulfanyl)-3-nitrobenzylidene]-2-thioxo-1,3-thiazolidin-4-one
-
-
(5Z)-5-[4-[(4-bromophenyl)sulfanyl]-3-nitrobenzylidene]-2-thioxo-1,3-thiazolidin-4-one
-
-
(5Z)-5-[4-[(4-chlorophenyl)sulfanyl]-3-nitrobenzylidene]-2-thioxo-1,3-thiazolidin-4-one
-
-
(5Z)-5-[4-[(4-fluorobenzyl)oxy]-3-nitrobenzylidene]-2-thioxo-1,3-thiazolidin-4-one
-
-
(5Z)-5-[4-[(4-fluorobenzyl)oxy]benzylidene]-2-thioxo-1,3-thiazolidin-4-one
-
-
(5Z)-5-[4-[(4-methylphenyl)sulfanyl]-3-nitrobenzylidene]-3-(prop-2-en-1-yl)-2-thioxo-1,3-thiazolidin-4-one
-
-
(9-adenylmethylcarbonyl)-4-aminobutyl triphosphate
-
fully competitive with respect to the nucleotide substrate dTTP, inhibits wild-type enzyme and mutant enzyme Y505A
(biphenylcarbonyl)-4-oxobutyl triphosphate
-
fully competitive with respect to the nucleotide substrate dTTP, inhibits wild-type enzyme and mutant enzyme Y505A; inhibits wild-type enzyme and mutant enzyme Y505A
(E)-enedione
-
-
1,2,3,4-tetrahydro-5-methoxynaphthalene-1,4-diol
-
i.e. nodulisporol. Strong inhibition of pol lambda, but does not influence the activities of mammalian pols alpha to kappa, and shows no effect on the activities of plant pols alpha and beta, prokaryotic pols, and other DNA metabolic enzymes such as calf terminal deoxynucleotidyl transferase, human immunodeficiency virus type-1 (HIV-1) reverse transcriptase, human telomerase, T7 RNA polymerase, and bovine deoxyribonuclease I
1,3-bis[2-chloroethyl]-2-nitrosourea
-
-
1-deoxyrubralactone
-
potent inhibitor of isozymes pol kappa, pol lambda, pol iota, and pol eta, but does not inhibit DNA polymerase isozymes pol delta, pol epsilon, and pol gamma
10-epi-pyragonicin
-
-
19-epi-jimenezin
-
-
2',3'-dideoxy-ATP
-
poor inhibitor
2-(p-n-Butylanilino)-2'-deoxyadenosine 5'-triphosphate
2-butylanilino-dATP
-
potent and highly selective inhibitor
2-methoxy-4-[(Z)-(4-oxo-2-thioxo-1,3-thiazolidin-5-ylidene)methyl]phenyl 2,4-dichlorobenzoate
-
-
2-methoxy-4-[(Z)-(4-oxo-2-thioxo-1,3-thiazolidin-5-ylidene)methyl]phenyl 3-bromobenzoate
-
-
2-thiomethyl-6-phenyl-4-(4'-hydroxybutyl)-1,2,4,-triazole (5,1-C)(1,2,4)triazine-7-one triphosphate
-
fully competitive with respect to the nucleotide substrate dTTP; fully competitive with respect to the nucleotide substrate dTTP, inhibits wild-type enzyme and mutant enzyme Y505A
3,4-dihydro-4-hydroxy-8-methoxynaphthalen-1(2H)-one
-
i.e. nodulisporone. Strong inhibition of pol lambda, but does not influence the activities of mammalian pols alpha to kappa, and shows no effect on the activities of plant pols alpha and beta, prokaryotic pols, and other DNA metabolic enzymes such as calf terminal deoxynucleotidyl transferase, human immunodeficiency virus type-1 (HIV-1) reverse transcriptase, human telomerase, T7 RNA polymerase, and bovine deoxyribonuclease I
3,5-dimethyl-8-methoxy-3,4-dihydroisocoumarin
-
-
4-chloromercuribenzoic acid
-
-
4-chlorophenyl 2,4-dinitrophenyl sulfide
-
-
4-hydroxy-5-methyl-3-tetradecyl-dihydrofuran-2(3H)-one
-
-
4-[(4-methylphenyl)sulfanyl]-3-nitrobenzaldehyde
-
-
4-[(Z)-(2,4-dioxo-1,3-thiazolidin-5-ylidene)methyl]-2-methoxyphenyl 3-bromobenzoate
-
-
4-[(Z)-(4-oxo-2-thioxo-1,3-thiazolidin-5-ylidene)methyl]phenyl 2-chlorobenzoate
-
-
5-(5-nitro-2-[[6-(trifluoromethyl)pyridin-3-yl]sulfanyl]benzylidene)-2-thioxo-1,3-thiazolidin-4-one
-
-
5-[2-(cyclohexylsulfanyl)-5-nitrobenzylidene]-2-thioxo-1,3-thiazolidin-4-one
-
-
5-[2-[(4-chlorophenyl)sulfanyl]-5-nitrobenzylidene]-2-thioxo-1,3-thiazolidin-4-one
-
-
5-[2-[(4-methylphenyl)sulfanyl]-5-nitrobenzylidene]-2-thioxo-1,3-thiazolidin-4-one
-
-
adefovir diphosphate
-
-
Allicin
-
-
alliin
-
-
aphidicolin
Ara-ATP
-
-
aurintricarboxylic acid
-
potent nanomolar inhibitor of pol beta, pol iota, and pol zeta
Ca2+
-
polymerase alpha
Carbonyldiphosphonate
cholesterol hemisuccinate
-
pol beta; pol iota; pol kappa; pol lambda
cloretazine
-
i.e. 1,2-bis[methylsulfonyl]-1-[2-chloroethyl]-2-[(methylamino)carbonyl]hydrazine
diallyl pentasulfide
-
-
diallyl tetrasulfide
-
-
diallyl trisulfide
-
-
Dideoxynucleoside 5'-triphosphate
Dideoxythymidine triphosphate
dimethyl sulfoxide
-
stimulates DNA polymerase alpha and delta, inhibits human DNA polymerase epsilon
ellagic acid
-
potent nanomolar inhibitor of pol beta, pol iota, and pol zeta
epigallocatechin gallate
-
-
gemcitabine
-
i.e. 2'-deoxy-2',2'-difluorocytidine, when gemcitabine is encountered as a template base DNA polymerase gamma pauses at the lesion and one downstream position but eventually elongates the primer to full-length product, these pauses are because of a 1000fold decrease in nucleotide incorporation efficiency
hymenoic acid
-
specific inhibitor of human DNA polymerase lambda, non-competitive inhibition with respect to both the DNA template-primer and the dNTP, i.e trans-4-[(1'E,5'S)-5'-carboxy-1'-methyl-1'-hexenyl]cyclohexanecarboxylic acid, does not influence the activities of the other mammalian pols alpha, gamma, delta, epsilon, eta, iota, and kappa, and also shows no effect even on the activity of pol beta
jimenezin
-
-
lamivudine triphosphate
-
-
Li+
-
polymerase alpha
Mg2+
-
DNA polymerase alpha: free Mg2+ competes with primer for enzyme binding, dramatic inhibition at Mg2+ concentration above the optimum
muconin
-
-
N-(2,4-dinitro-5-fluorophenyl)-2-aminoethyl triphosphate
-
inhibition of mutant enzyme Y505A, inactive against wild-type enzyme
N-(2,4-dinitro-5-fluorophenyl)-4-aminobutyl triphosphate
-
fully competitive with respect to the nucleotide substrate dTTP, inhibits wild-type enzyme and mutant enzyme Y505A
N-(2,4-dinitro-5-imidazolylphenyl)-2-aminoethyl triphosphate
-
inhibition of mutant enzyme Y505A, inactive against wild-type enzyme
N-(2,4-dinitro-5-imidazolylphenyl)-4-aminobutyl triphosphate
-
inhibition of mutant enzyme Y505A, inactive against wild-type enzyme
N-(2,4-dinitrophenyl)-4-aminobutyl triphosphate
-
fully competitive with respect to the nucleotide substrate dTTP, inhibits wild-type enzyme and mutant enzyme Y505A
N-(benzyloxycarbonyl)-4-aminobutyl triphosphate
-
fully competitive with respect to the nucleotide substrate dTTP, inhibits wild-type enzyme and mutant enzyme Y505A; inhibits wild-type enzyme and mutant enzyme Y505A
N-ethylmaleimide
N-[6-N-(2,4-dinitrophenyl)aminohexanoyl]-2-aminoethyl triphosphate
-
inhibition of mutant enzyme Y505A, inactive against wild-type enzyme
N2-(p-n-butylphenyl)-2'-deoxyguanosine 5'-triphosphate
-
inhibition of DNA polymerase alpha at 100fold; lower concentration than DNA polymerase delta
N2-CH2(6-benzo[a]pyrenyl)guanine
-
severely blocks activity
-
N2-CH2(9-anthracenyl)guanine
-
severely blocks activity, frequencies of dATP misinsertion and extension beyond mispairs are proportionally increased, great decrease in pre-steady-state kinetic burst rate. 2.6fold decrease in dCTP binding affinity and increased DNA substrate binding affinity
-
NSC-666715
-
potent, small molecular weight inhibitor of DNA polymerase beta, blocks Pol-beta-directed single-nucleotide- and long-patch-base excisison repair
Oosporein
-
50% inhibition at 0.61 mM
pamoic acid
-
pol beta inhibitor
plakevulin A
-
-
pyragonicin
-
-
pyranicin
-
potent inhibitor
rhodanines
-
most potent inhibitors for DNA Pol lambda, they are up to 10times less active against the highly similar DNA polymerase beta, structure-activity relationships, overview. 5-Arylidene-2,4-thiazolidinediones are class I rhodanines, rhodanine class II has members of carbohydrazides, and class III contains a common 2,4-pentadione substructure element
-
Salt
-
concentrations above 50 mM inhibit: human KB cell polymerase alpha
single-stranded DNA
-
inhibition of polymerase alpha, competitive with respect to activated DNA substrate
stavudine triphosphate
-
-
talaroflavone
-
-
tenofovir diphosphate
-
-
untenone A
-
-
vitamin K3
-
more than 80% inhibition of pol gamma at 0.03 mM, does not affect other human DNA polymerase isozymes
zidovudine triphosphate
-
-
[3-[(Z)-(4-oxo-2-thioxo-1,3-thiazolidin-5-ylidene)methyl]phenoxy]acetic acid
-
-
additional information
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
dimethyl sulfoxide
-
stimulates DNA polymerase alpha and delta, inhibits human DNA polymerase epsilon
FEN1
-
stimulates strand displacement activity, DEN1 processes nicked DNA, thus removing a barrier to Pol lambda DNA synthesis. It results in a one-nucleotide gapped DNA molecule that is a favorite substrate of Pol lambda
-
PCNA
-
stimulates strand displacement activity
-
Polymerase alpha accessory factors
-
overview
-
Proliferating cell nuclear antigen
-
Replication factor A
-
RF-A, multisubunit single-stranded DNA-binding protein, functions as an auxiliary protein for both polymerases alpha and delta, required for initiation and elongation stages of in vitro SV40 DNA replication
-
Replication factor C
-
RF-C, multisubunit protein complex with primer/template binding and DNA-dependent ATPase activity, has a profound effect on leading-strand DNA synthesis
-
spermidine
-
up to 10 mM, polymerase beta
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00249 - 0.00792
dCTP
0.00118
deoxynucleoside triphosphate
-
DNA polymerase lambda, in 50 mM Tris-HCl (pH 7.5), 1 mM dithiothreitol, 50% (v/v) glycerol, and 0.1 mM EDTA, at 37°C
0.175 - 0.211
dTTP
additional information
additional information
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
additional information
Homo sapiens
-
-
-
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0028 - 0.0038
(9-adenylmethylcarbonyl)-4-aminobutyl triphosphate
0.0004 - 0.0048
(biphenylcarbonyl)-4-oxobutyl triphosphate
0.0188
2',3'-dideoxy-ATP
-
-
0.001
2-butylanilino-dATP
-
-
0.0018 - 0.068
2-thiomethyl-6-phenyl-4-(4'-hydroxybutyl)-1,2,4,-triazole (5,1-C)(1,2,4)triazine-7-one triphosphate
0.00271
adefovir diphosphate
-
-
0.00065
aphidicolin
-
-
0.0506
lamivudine triphosphate
-
-
0.0144
N-(2,4-dinitro-5-fluorophenyl)-2-aminoethyl triphosphate
-
mutant enzyme Y505A
0.0031 - 0.0065
N-(2,4-dinitro-5-fluorophenyl)-4-aminobutyl triphosphate
0.0005
N-(2,4-dinitro-5-imidazolylphenyl)-2-aminoethyl triphosphate
-
mutant enzyme Y505A
0.0154
N-(2,4-dinitro-5-imidazolylphenyl)-4-aminobutyl triphosphate
-
mutant enzyme Y505A
0.0012 - 0.0035
N-(2,4-dinitrophenyl)-4-aminobutyl triphosphate
0.0016 - 0.07
N-(benzyloxycarbonyl)-4-aminobutyl triphosphate
0.0091
N-[6-N-(2,4-dinitrophenyl)aminohexanoyl]-2-aminoethyl triphosphate
-
mutant enzyme Y505A
0.0569
stavudine triphosphate
-
-
0.00618
tenofovir diphosphate
-
-
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.011 - 0.1
(2E)-2-(3-nitro-4-[[6-(trifluoromethyl)pyridin-3-yl]sulfanyl]benzylidene)-5-thioxo-1,3-thiazolidin-4-one
0.0083 - 0.08
(5Z)-5-[3-bromo-4-[(4-fluorophenyl)sulfanyl]benzylidene]-2-thioxo-1,3-thiazolidin-4-one
0.0093 - 0.1
(5Z)-5-[4-(4-methylphenoxy)-3-nitrobenzylidene]-2-thioxo-1,3-thiazolidin-4-one
0.0059 - 0.0644
(5Z)-5-[4-[(4-fluorobenzyl)oxy]benzylidene]-2-thioxo-1,3-thiazolidin-4-one
0.0037 - 0.0088
(E)-enedione
0.092 - 0.307
1,2-bis[methylsulfonyl]-1-[(methylamino)carbonyl]hydrazine
0.271 - 1.435
1,3-bis[2-chloroethyl]-2-nitrosourea
0.0598
1-deoxyrubralactone
Homo sapiens
-
in 50 mM Tris-HCl (pH 7.5) containing 1 mM dithiothreitol, 50% (v/v) glycerol, and 0.1 mM EDTA, at 37°C
0.049
3,5-dimethyl-8-methoxy-3,4-dihydroisocoumarin
Homo sapiens
-
-
0.0124 - 0.0888
4-[(Z)-(4-oxo-2-thioxo-1,3-thiazolidin-5-ylidene)methyl]phenyl 2-chlorobenzoate
0.0081 - 0.0429
5-[2-[(4-chlorophenyl)sulfanyl]-5-nitrobenzylidene]-2-thioxo-1,3-thiazolidin-4-one
0.01 - 0.0455
5-[2-[(4-methylphenyl)sulfanyl]-5-nitrobenzylidene]-2-thioxo-1,3-thiazolidin-4-one
0.0063 - 0.148
cholesterol hemisuccinate
0.074 - 0.387
cloretazine
0.146
diallyl trisulfide
Homo sapiens
-
DNA polymerase lambda, in 50 mM Tris-HCl (pH 7.5), 1 mM dithiothreitol, 50% (v/v) glycerol, and 0.1 mM EDTA, at 37°C
0.0038
epigallocatechin gallate
Homo sapiens
-
pH and temperature not specified in the publication
0.0917
hymenoic acid
Homo sapiens
-
-
0.0049 - 0.06
pamoic acid
0.066 - 0.179
plakevulin A
0.0023 - 0.0158
pyranicin
0.0043 - 0.057
untenone A
0.006
vitamin K3
Homo sapiens
-
isozyme pol gamma, in 50 mM Tris-HCl (pH 7.5) containing 1 mM dithiothreitol, 50% (v/v) glycerol, and 0.1 mM EDTA, at 37°C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.968
-
polymerase beta
3.43
-
polymerase alpha
5.33
-
-
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.5 - 7.9
-
assay at
7.5 - 8.5
-
KB cell DNA polymerase alpha
8
-
assay at
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.2 - 9
-
pH 7.2: about 65% of maximal activity, pH 9.0: about 40% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
22
-
assay at room temperature
37
-
assay at
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5
-
polymerase alpha
5.6 - 5.9
-
polymerase C, sucrose gradient, ampholytes pH 3-11
8.5
-
polymerase beta
9.3
-
polymerase N1, sucrose gradient, ampholytes pH 3-11
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
additional information
human DNA polymerase eta expression is detected in most tissues except for very low or undetectable levels in peripheral lymphocytes, fetal spleen, and adult muscle
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
78410
estimated from amino acid sequence
40000
-
vertebrates, DNA polymerase beta
45000
-
1 * 45000 + 1 * 60000, SDS-PAGE, mitochondrial enzyme
48000
-
1 * 125000 catalytic subunit + 1 * 48000 subunit of unknown function, DNA polymerase delta
54000
-
1 * 54000 + 1 * 140000
60000
-
1 * 45000 + 1 * 60000, SDS-PAGE, mitochondrial enzyme
66000
-
1 * 66000 + 1 * 76000, DNA polymerase alpha, SDS-PAGE
76000
-
1 * 66000 + 1 * 76000, DNA polymerase alpha, SDS-PAGE
77100
-
N-terminally His6-tagged enzyme fragment Pol ny-77, gel filtration
87000
-
1 * 155000-175000 + 1 * 87000, SDS-PAGE, polymerase alpha
100000 - 110000
-
mitochondrial enzyme
100000
-
full-length Pol ny
110000
120000
125000
-
1 * 125000 catalytic subunit + 1 * 48000 subunit of unknown function, DNA polymerase delta
140000
170000
190000
215000
-
DNA polymerase epsilon
265000 - 280000
-
dimeric form, polymerase alpha
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
-
x * 140000, catalytic subunit, + 55000, accessory subunit, SDS-PAGE
monomer
tetramer
-
-
additional information
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
pol beta-DNA complexes dGTP(beta-gamma CF2) and dGTP(beta-gamma CH2) are crystallized by sitting-drop vapor diffusion
-
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25
-
stable for 3 h in an optimized reaction buffer
37
-
lifetime below 20 min
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
0°C, in concentrated form in the presence of sucrose and potassium phosphate, half-life: 1.5 months, human KB cell DNA polymerase alpha
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
3 non-mitochondrial enzymes from KB cells: C, N1 and N2
-
DNA polymerase alpha
-
native Pol delta from UV-treated HeLa cells by immunoaffinity chromatography using anti-p125-agarose, recombinant Pol delta subunits from Sf9 insect cells by immunoaffinity and anion exchange chromatography, recombinant His-tagged or GST-tagged subunits from Escherichia coli by nickel affinity and glutathione affinity chromatography, respectively
-
nickel affinity column chromatography and SP column chromatography
-
polymerase delta
-
polymerase gamma
-
recombinant
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recombinant C-terminally His6-tagged full-length p55 subunit from Escherichia coli strain BL21(DE3) to homogeneity, by nickel affinity chromatography, recombinant N-terminally His6-tagged wild-type enzyme and mutant R946C from Sf9 insect cells by nickel affinity chromatography, gel filtration, and anion exchange chromatography
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recombinant refolded N-terminally His6-tagged enzyme fragment Pol ny-77 by gel filtration, dialysis, and heparin affinity chromatography, recombinant soluble His6-tagged enzyme fragment Pol ny-77 from Escherichia coli by nickel affinity chrmatography
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
catalytic subunit of DNA polymerase delta
catalytic subunits of wild-type and exonuclease-deficient mutant
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expressed in Escherichia coli BL21(DE3) cells
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expressed in Sf9 cells
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expression in Escherichia coli
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expression of His-tagged proteins of His-p125, His-p50, His-p68, and Hisp12, in Escherichia coli strain BL21DE3(plys), expression of GST-tagged p12 using pGEX-5X-3 vector in Escherichia coli strain BL21DE3(plys), reconstitution of Pol delta complexes in insect cells infected with a single baculovirus into which one or more subunits are assembled, allowing efficient expression of the tetrameric Pol delta holoenzyme, the p125/p50 core dimer, the core+p68 trimer and the core+p12 trimer, as well as the p125 catalytic subunit
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gene POLN, expression of N-terminally His6-tagged enzyme fragment Pol ny-77 in Escherichia coli in inclusion bodies
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overexpressed in Escherichia coli
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overview: genetic structure and predicted functional domains
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the catalytic subunit is encoded by gene POLG at chromosomal locus 15q2, the accessory subunit is encoded by gene POLG2 at chromosomal locus 17q24.1, expression of polymerase point mutations combined with the D198A/E200A exonuclease mutation, the mutant is also lacking the mitochondrial targeting sequence, expression of N-terminally His6-tagged wild-type enzyme and mutant R946C in Spodoptera frugiperda Sf9 cells, expression of His6-tagged full-length p55 subunit without its mitochondrial targeting sequence in Escherichia coli strain BL21(DE3)
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
DNA polymerase eta expression is induced by cisplatin, mRNA expression can be up-regulated in a p53-dependent manner following ionizing radiation or camptothecin treatment
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
A467T
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naturally occuring mutation, the mutation is the most common POLG mutation and has been found to be associated with all of the disease symptoms analyzed. The A467T pol gamma possesses only 4% of the wild-type DNA polymerase activity and is compromised for its ability to interact with the p55 accessory subunit
A957S
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naturally occuring mutation, involved in autosomal dominant progressive external ophthalmoplegia, the mutation is associated with motiif B in the active site
D115A/E116A
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catalytically inactive
D198A/E200A
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site-directed mutagenesis in the exonuclease domain resulting in loss of exonuclease activity
E1143G
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naturally occuring mutation, that is a frequent cause of ataxia-neuropathy syndrome, and found in 4% of European populations
E200A
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exonuclease-deficient mutant
F506G
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inactive mutant enzyme
F506R
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complete loss of de novo DNA synthesis
G848S
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naturally occuring mutation, involved in Alpers syndrome, the mutant shows compromised DNA binding ability that affects its overall functional efficiency
G923D
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naturally occuring mutation, involved in autosomal dominant progressive external ophthalmoplegia, the mutation is associated with motiif B in the active site
L606G
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site-directed mutagenesis, the mutant shows increased polymerase activity and slightly reduced exonuclease activity compared to the wild-type enzyme, mutant pol delta L606G is highly error prone, incorporating single noncomplementary nucleotides at a high frequency during DNA synthesis
L606K
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site-directed mutagenesis, the mutant shows increased polymerase activity and reduced exonuclease activity compared to the wild-type enzyme, mutant pol delta L606K is extremely accurate, with a higher fidelity of single nucleotide incorporation by the active site than that of wild-type pol delta, it does not catalyze detectable nucleotide mis-insertion even with nucleotide concentrations as high as 4 mM, but pol delta L606K mutant is impaired in the bypass of DNA adducts
R852C
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naturally occuring mutation, involved in Alpers syndrome, the mutant shows compromised DNA binding ability that affects its overall functional efficiency
R853Q
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naturally occuring mutation, involved in Alpers syndrome, the mutant shows compromised DNA binding ability that affects its overall functional efficiency
R943H
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naturally occuring mutation, involved in autosomal dominant progressive external ophthalmoplegia, the mutation is associated with motiif B in the active site, the mutant retains less than 1% of the wild-type polymerase activity and displays a severe decrease in processivity
T851A
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naturally occuring mutation, involved in Alpers syndrome, the mutant shows compromised DNA binding ability that affects its overall functional efficiency
W748S
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naturally occuring mutation that has intrinsic lower polymerase activity as well as a demonstrated lower affinity for DNA compared to the wild-type enzyme
W748S/E1143G
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naturally occuring mutation, the E1143G single-nucleotide polymorphism can modulate the deleterious effect of the W748S mutation
Y955C
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naturally occuring mutation, involved in autosomal dominant progressive external ophthalmoplegia, the mutation is associated with motiif B in the active site, the mutant retains less than 1% of the wild-type polymerase activity and displays a severe decrease in processivity, the mutation increases nucleotide misinsertion replication errors 10-100 fold in the absence of exonucleolytic proofreading
Renatured/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
use of high hydrostatic pressure to refold recombinant N-terminally His6-tagged enzyme fragment Pol ny-77 from inclusion bodies after expression in Escherichia coli, in soluble and active form. The refolded Pol ny fragment has properties comparable to those of the small amount of Pol m that is purified from the soluble fraction, method evaluation
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
drug development
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DNA polymerases are promising drug targets for the treatment of cancer
medicine