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Information on EC 3.2.2.27 - uracil-DNA glycosylase and Organism(s) Thermus thermophilus and UniProt Accession Q5SKC5
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3.2.2.27 uracil-DNA glycosylaseIUBMB Comments
Uracil-DNA glycosylases are widespread enzymes that are found in all living organisms. EC 3.2.2.27 and double-stranded uracil-DNA glycosylase (EC 3.2.2.28) form a central part of the DNA-repair machinery since they initiate the DNA base-excision repair pathway by hydrolysing the N-glycosidic bond between uracil and the deoxyribose sugar thereby catalysing the removal of mis-incorporated uracil from DNA.
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Word Map
- 3.2.2.27
-
deamination
- cytosine
- glycosylases
- endonuclease
- thymine
-
abasic
-
misincorporation
- ung2
-
uracil-containing
- duplex
-
hypermutation
- dump
- deoxyuridine
-
activation-induced
-
excises
- mispairs
-
base-excision
- dutpase
- 8-oxoguanine
-
apyrimidinic
- 5-methylcytosine
- 5-hydroxymethyluracil
-
g-quadruplexes
-
transversions
- ape1
-
thymine-dna
- formamidopyrimidine
- 5-hydroxyuracil
-
short-patch
- xrcc1
-
apurinic
-
class-switching
- 3,n4-ethenocytosine
-
translesion
-
post-replicative
- ap-endonuclease
-
aid-induced
- apobec3g
- drug development
- molecular biology
- analysis
- medicine
The taxonomic range for the selected organisms is: Thermus thermophilus
The enzyme appears in selected viruses and cellular organisms
The enzyme appears in selected viruses and cellular organisms
Reaction Schemes
Hydrolyses single-stranded DNA or mismatched double-stranded DNA and polynucleotides, releasing free uracil
Synonyms
uracil-dna glycosylase, smug1, dna n-glycosylase, ung-1, ul114, uracil dna-glycosylase, uracil-dna n-glycosylase, uracil dna glycosylase 2, thd1p, mjudg, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
UDG
additional information
additional information
the enzyme is an UDG family 5 enzyme belonging to the UDG superfamily
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Hydrolyses single-stranded DNA or mismatched double-stranded DNA and polynucleotides, releasing free uracil
catalytic reaction mechanism, the active side comprises the GEGPG motif, residues 40-44, the side-chain of Arg161 in family 4 TthUDG might play a role in binding AP-DNA after catalysis
Q7WYV4
Hydrolyses single-stranded DNA or mismatched double-stranded DNA and polynucleotides, releasing free uracil
a conserved asparagine residue acts as a ligand to the catalytic water molecule, and another water molecule acts as a barrier during substrate recognition, slide-in mechanism for initial damage recognition, catalytic reaction mechanism, overview
SYSTEMATIC NAME
IUBMB Comments
uracil-DNA deoxyribohydrolase (uracil-releasing)
Uracil-DNA glycosylases are widespread enzymes that are found in all living organisms. EC 3.2.2.27 and double-stranded uracil-DNA glycosylase (EC 3.2.2.28) form a central part of the DNA-repair machinery since they initiate the DNA base-excision repair pathway by hydrolysing the N-glycosidic bond between uracil and the deoxyribose sugar thereby catalysing the removal of mis-incorporated uracil from DNA.
CAS REGISTRY NUMBER
COMMENTARY
59088-21-0
cf. EC 3.2.2.28
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
double stranded DNA containing G-U mismatch + H2O
uracil + double-stranded DNA with abasic site
-
-
-
?
uracil-containing single-stranded DNA + H2O
uracil + single-stranded DNA with abasic site
-
-
-
?
uracil-mismatched double-stranded DNA + H2O
uracil + double-stranded DNA with abasic site
Q7WYV4
-
-
-
?
uracil-mismatched double-stranded DNA + H2O
uracil + double-stranded DNA with abasic site
Q7WYV4
UDG removes uracil from DNA to initiate DNA base excision repair, Thermus thermophilus UDG processes both single-stranded and double-stranded DNA containing uracil, regardless of opposing base, but does not process G-T mismatched DNA, nor does it possess AP endonuclease activity, uracil bases in U-A mismatches are excised less efficiently, due to the stability of that particular base-pair. The UDG possesses a [4Fe-4S] cluster, distant from the active site, which interacts with loop structures and is unessential to the activity but necessary for stabilizing the loop structures. Uracil recognition mechanism, overview
-
-
?
uracil-mismatched single-stranded DNA + H2O
uracil + single-stranded DNA with abasic site
Q7WYV4
-
-
-
?
uracil-mismatched single-stranded DNA + H2O
uracil + single-stranded DNA with abasic site
Q7WYV4
UDG removes uracil from DNA to initiate DNA base excision repair, Thermus thermophilus UDG processes both single-stranded and double-stranded DNA containing uracil, regardless of opposing base, but does not process G-T mismatched DNA, nor does it possess AP endonuclease activity, uracil bases in U-A mismatches are excised less efficiently, due to the stability of that particular base-pair. The UDG possesses a [4Fe-4S] cluster, distant from the active site, which interacts with loop structures and is unessential to the activity but necessary for stabilizing the loop structures. Uracil recognition mechanism, overview
-
-
?
uracil-mismatched double-stranded DNA + H2O
uracil + double-stranded DNA with abasic site
-
-
-
?
uracil-mismatched double-stranded DNA + H2O
uracil + double-stranded DNA with abasic site
-
-
-
?
additional information
?
-
Q7WYV4
UDG is an essential enzyme for maintaining the integrity of genomic information, it is the first enzyme of a base excision repair, BER, pathway that corrects uracil lesions. TthUDG specifically recognizes uracil that is flipped out from double-stranded DNA, in a manner similar to that of the family 1 human UDG, rather than binding to the guanine base of the complementary strand in mismatched DNA, as does the family 2 Escherichia coli MUG
-
-
?
additional information
?
-
UDG removes uracil generated by the deamination of cytosine or misincorporation of deoxyuridine monophosphate. The fifth UDG family lacks a polar residue in the active-site motif, which mediates the hydrolysis of the glycosidic bond by activation of a water molecule in UDG families 1 to 4
-
-
?
additional information
?
-
family 5 UDGB in complex with rAP-G DNA and rAP-A DNA, substrate specificity and binding structure analysis, modelling, overview
-
-
?
additional information
?
-
the enzyme does not show any detectable activity on other deaminated bases than uracil
-
-
?
additional information
?
-
-
the enzyme does not show any detectable activity on other deaminated bases than uracil
-
-
?
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
uracil-mismatched double-stranded DNA + H2O
uracil + double-stranded DNA with abasic site
Q7WYV4
-
-
-
?
uracil-mismatched single-stranded DNA + H2O
uracil + single-stranded DNA with abasic site
Q7WYV4
-
-
-
?
uracil-containing single-stranded DNA + H2O
uracil + single-stranded DNA with abasic site
-
-
-
?
uracil-mismatched double-stranded DNA + H2O
uracil + double-stranded DNA with abasic site
-
-
-
?
uracil-mismatched double-stranded DNA + H2O
uracil + double-stranded DNA with abasic site
-
-
-
?
additional information
?
-
Q7WYV4
UDG is an essential enzyme for maintaining the integrity of genomic information, it is the first enzyme of a base excision repair, BER, pathway that corrects uracil lesions. TthUDG specifically recognizes uracil that is flipped out from double-stranded DNA, in a manner similar to that of the family 1 human UDG, rather than binding to the guanine base of the complementary strand in mismatched DNA, as does the family 2 Escherichia coli MUG
-
-
?
additional information
?
-
UDG removes uracil generated by the deamination of cytosine or misincorporation of deoxyuridine monophosphate. The fifth UDG family lacks a polar residue in the active-site motif, which mediates the hydrolysis of the glycosidic bond by activation of a water molecule in UDG families 1 to 4
-
-
?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Fe2+
Q7WYV4
the UDG possesses a [4Fe-4S] cluster, distant from the active site, which interacts with loop structures and is unessential to the activity but necessary for stabilizing the loop structures
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.001
uracil-mismatched single-stranded DNA
Q7WYV4
pH 7.5, 37°C, family 4 UDG
-
0.00017
double stranded DNA containing G-U mismatch
mutant enzyme E41Q/G42D, at pH 7.6 and 60°C
-
0.00074
double stranded DNA containing G-U mismatch
mutant enzyme N89A, at pH 7.6 and 60°C
-
0.00076
double stranded DNA containing G-U mismatch
mutant enzyme H155S, at pH 7.6 and 60°C
-
0.00097
double stranded DNA containing G-U mismatch
wild type enzyme, at pH 7.6 and 60°C
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.17
uracil-mismatched single-stranded DNA
Q7WYV4
pH 7.5, 37°C, family 4 UDG
-
0.000072
double stranded DNA containing G-U mismatch
mutant enzyme H155S, at pH 7.6 and 60°C
-
0.0075
double stranded DNA containing G-U mismatch
mutant enzyme E41Q/G42D, at pH 7.6 and 60°C
-
0.025
double stranded DNA containing G-U mismatch
mutant enzyme N89A, at pH 7.6 and 60°C
-
0.23
double stranded DNA containing G-U mismatch
wild type enzyme, at pH 7.6 and 60°C
-
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0092
double stranded DNA containing G-U mismatch
mutant enzyme H155S, at pH 7.6 and 60°C
-
35
double stranded DNA containing G-U mismatch
mutant enzyme N89A, at pH 7.6 and 60°C
-
45
double stranded DNA containing G-U mismatch
mutant enzyme E41Q/G42D, at pH 7.6 and 60°C
-
240
double stranded DNA containing G-U mismatch
wild type enzyme, at pH 7.6 and 60°C
-
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
additional information
Q7WYV4
the UDG possesses a [4Fe-4S] cluster, distant from the active site, which interacts with loop structures and is unessential to the activity but necessary for stabilizing the loop structures, salt-bridges and ion pairs on the molecular surface and the presence of proline on loops and turns contribute to the enzyme's thermostability
additional information
family 5 UDGB secondary structure elements, overview
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
family 4 UDG in complex with uracil, hanging drop vapor diffusion method, mixing of 0.002 ml of 13 mg/ml of selenomethionyl protein solution with 0.002 ml of 1.2-1.5 M ammonium sulfate, 25% v/v glycerol, and 75 mM Tris-HCl, pH 8.5, and equilibration against 0.3 ml of the reservoir solution at 4°C, X-ray diffraction structure determination and analysis at 1.5 A resolution
Q7WYV4
recombinant family 5 UDGB, as free protein or selenomethionine-labeled protein, or in complex with rAP-G DNA and rAP-A DNA, hanging drop vapor diffusion method, 0.001 ml drops of 11 mg/ml TtUDGB are mixed with 0.001 ml of 9-13% v/v PEG 3350, 0.2 M ammonium acetate, 5% v/v glycerol and 0.1 M MES, pH 6.5, and equilibrated against 0.5 ml of the reservoir solution at 20°C. X-ray diffraction structure determination and analysis at 1.45-2.1 A resolution, molecular replacement
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
D75A
the D75A mutant shows low enzymatic activity for the removal of uracil from U-G or thymine from T-G. However, the mutant can distinguish between the C5-hydrogen and the C5-methyl group
E41A/G42D
the mutant shows severely reduced activity compared to the wild type enzyme
E41Q
the mutant shows 888fold reduced activity compared to the wild type enzyme
E41Q/G42D
the mutant shows 5.3fold reduced activity compared to the wild type enzyme
E47A
the mutant shows severely reduced activity compared to the wild type enzyme
F54A
the mutant shows severely reduced activity compared to the wild type enzyme
G42D
the mutant shows 89fold reduced activity compared to the wild type enzyme
H155S
the mutant shows reduced activity compared to the wild type enzyme
N80A
the mutant shows severely reduced activity compared to the wild type enzyme
N89A
the mutant shows reduced activity compared to the wild type enzyme
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
Q7WYV4
salt-bridges and ion pairs on the molecular surface and the presence of proline on loops and turns contribute to the enzyme's thermostability
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant wild-type and selenomethionine-labeled enzymes from Escherichia coli strain BL21(DE3) by hydrophobic interaction and anion exchange chromatography, followed by hydroxyapatite adsorption chromatography and gel filtration
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
gene TtUDGB, expression of wild-type and selenomethionine-labeled enzymes in Escherichia coli strain BL21(DE3)
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Hoseki, J.; Okamoto, A.; Masui, R.; Shibata, T.; Inoue, Y.; Yokoyama, S.; Kuramitsu, S.
Crystal structure of a family 4 uracil-DNA glycosylase from Thermus thermophilus HB8
J. Mol. Biol.
333
515-526
2003
Thermus thermophilus (Q7WYV4), Thermus thermophilus HB8 / ATCC 27634 / DSM 579 (Q7WYV4)
Kosaka, H.; Hoseki, J.; Nakagawa, N.; Kuramitsu, S.; Masui, R.
Crystal structure of family 5 uracil-DNA glycosylase bound to DNA
J. Mol. Biol.
373
839-850
2007
Thermus thermophilus (Q5SJG5), Thermus thermophilus HB8 / ATCC 27634 / DSM 579 (Q5SJG5)
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