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Information on EC 3.2.2.27 - uracil-DNA glycosylase and Organism(s) Escherichia coli and UniProt Accession P12295
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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: Escherichia coli
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
additional information
-
the enzyme belongs to the family I uracil-DNA glycosylases
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Hydrolyses single-stranded DNA or mismatched double-stranded DNA and polynucleotides, releasing free uracil
active site structure and catalytic 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
uracil-mismatched single-stranded DNA + H2O
uracil + single-stranded DNA with abasic site
family 2 mismatch-specific uracil DNA glycosylase (MUG) is known to exhibit glycosylase activity on three mismatched base pairs, T/U, G/U and C/U. Family 1 uracil N-glycosylase (UNG) is an extremely efficient enzyme that can remove uracil from any uracil-containing base pairs including the A/U base pair
-
-
?
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
family 2 mismatch-specific uracil DNA glycosylase (MUG) is known to exhibit glycosylase activity on three mismatched base pairs, T/U, G/U and C/U. Family 1 uracil N-glycosylase (UNG) is an extremely efficient enzyme that can remove uracil from any uracil-containing base pairs including the A/U base pair
-
-
?
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
-
substrate specificity, overview
-
-
?
uracil-mismatched double-stranded DNA + H2O
uracil + double-stranded DNA with abasic site
-
the enzyme initiates repair of uracil-DNA is achieved in a base-excision pathway
-
-
?
uracil-mismatched double-stranded DNA + H2O
uracil + double-stranded DNA with abasic site
-
the enzyme hydrolyses the N-glycosidic bond connecting the base to the deoxyribose sugar of the DNA backbone, releasing free uracil base and DNA containing an abasic site, as its products, substrate recognition by family-1 UDG, modelling, detailed overview
-
-
?
uracil-mismatched double-stranded DNA + H2O
uracil + double-stranded DNA with abasic site
-
Ung can utilize both double- and single-stranded substrates, preferring the latter
-
-
?
uracil-mismatched single-stranded DNA + H2O
uracil + single-stranded DNA with abasic site
-
-
-
-
?
uracil-mismatched single-stranded DNA + H2O
uracil + single-stranded DNA with abasic site
-
Ung catalyzes the removal of uracil from Ura-Cyt pairs in single-stranded long DNA consisting of identical repeated lesion-containing oligonucleotide units, constructed by ligation. Ung can utilize both double- and single-stranded substrates, preferring the latter
-
-
?
additional information
?
-
preparation of a set of model nucleosome substrates, of 154mer DNA, in which single thymidine residues are replaced with uracil at specific locations and a second set of nucleosomes in which uracils are randomly substituted for all thymidines. UDG efficiently removes uracil from internal locations in the nucleosome where the DNA backbone is oriented away from the surface of the histone octamer, without significant disruption of histone-DNA interactions. However, uracils at sites oriented toward the histone octamer surface are excised at much slower rates, consistent with a mechanism requiring spontaneous DNA unwrapping from the nucleosome. In contrast to the nucleosome core, UDG activity on DNA outside the core DNA region is similar to that of naked DNA, overview
-
-
?
additional information
?
-
Ung shows strict specificity for uracil excision activity
-
-
?
additional information
?
-
-
family-1 enzymes are active against uracil in ssDNA and dsDNA, and recognise uracil explicitly in an extrahelical conformation via a combination of protein and bound-water interactions. Extrahelical recognition requires an efficient process of substrate location by base-sampling probably by hopping or gliding along the DNA. Family-2 enzymes are mismatch specific and explicitly recognise the widowed guanine on the complementary strand rather than the extrahelical scissile pyrimidine. Although structures are not yet available for family-3/SMUG and family-4 enzymes, sequence analysis suggests similar overall folds, and identifies common active site motifs but with a surprising lack of conservation of catalytic residues between members of the super-family
-
-
?
additional information
?
-
-
Ung catalyzes the removal of uracil from Ura-Cyt pairs in single-stranded long DNA consisting of identical repeated lesion-containing oligonucleotide units, constructed by ligation. Ung can utilize both double- and single-stranded substrates, preferring the latter
-
-
?
additional information
?
-
-
UNG removes uracil from DNA, substrate recognition and catalytic reaction mechanism, short-range sliding is vital for extrahelical uracil trapping, intramolecular transfer mechanisms of the enzyme, detailed overview
-
-
?
additional information
?
-
-
uracil-DNA glycosylases are ubiquitously found enzymes that hydrolyze the N-glycosidic bond of deoxyuridine, generating from deamination of cytosine, in DNA, UNG enzymes specifically excise Ura bases from both double-stranded and single-stranded DNA with a slight preference for the latter substrate, and shows no activity against normal DNA bases or against uracil in RNA. As potentially mutagenic and deleterious for cell regulation, uracil must be removed from DNA
-
-
?
additional information
?
-
-
UNG hydrolyzes the N-glycosidic bond of deoxyuridine in DNA. It binds with appreciable affinity to any DNA, mainly due to the interactions with the charged backbone. Search for the lesion by UNG involves random sliding along DNA alternating with dissociation-association events and partial eversion of undamaged bases for initial sampling. DNA in the complex with UNG is highly distorted to allow the extrahelical recognition of uracil, mechanism of uracil search and recognition by UNG, overview
-
-
?
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 single-stranded DNA + H2O
uracil + single-stranded DNA with abasic site
family 2 mismatch-specific uracil DNA glycosylase (MUG) is known to exhibit glycosylase activity on three mismatched base pairs, T/U, G/U and C/U. Family 1 uracil N-glycosylase (UNG) is an extremely efficient enzyme that can remove uracil from any uracil-containing base pairs including the A/U base pair
-
-
?
uracil-containing single-stranded DNA + H2O
uracil + single-stranded DNA with abasic site
-
-
-
-
?
uracil-mismatched 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
family 2 mismatch-specific uracil DNA glycosylase (MUG) is known to exhibit glycosylase activity on three mismatched base pairs, T/U, G/U and C/U. Family 1 uracil N-glycosylase (UNG) is an extremely efficient enzyme that can remove uracil from any uracil-containing base pairs including the A/U base pair
-
-
?
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
-
the enzyme initiates repair of uracil-DNA is achieved in a base-excision pathway
-
-
?
additional information
?
-
-
family-1 enzymes are active against uracil in ssDNA and dsDNA, and recognise uracil explicitly in an extrahelical conformation via a combination of protein and bound-water interactions. Extrahelical recognition requires an efficient process of substrate location by base-sampling probably by hopping or gliding along the DNA. Family-2 enzymes are mismatch specific and explicitly recognise the widowed guanine on the complementary strand rather than the extrahelical scissile pyrimidine. Although structures are not yet available for family-3/SMUG and family-4 enzymes, sequence analysis suggests similar overall folds, and identifies common active site motifs but with a surprising lack of conservation of catalytic residues between members of the super-family
-
-
?
additional information
?
-
-
uracil-DNA glycosylases are ubiquitously found enzymes that hydrolyze the N-glycosidic bond of deoxyuridine, generating from deamination of cytosine, in DNA, UNG enzymes specifically excise Ura bases from both double-stranded and single-stranded DNA with a slight preference for the latter substrate, and shows no activity against normal DNA bases or against uracil in RNA. As potentially mutagenic and deleterious for cell regulation, uracil must be removed from DNA
-
-
?
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
DNA containing 1'-cyano-2'-deoxyuridine
-
CNdU, a UDG inhibitor whose respective nucleotide triphosphate is also a substrate for DNA polymerase
-
additional information
association of linker histone reduces activity of UDG at selected sites near where the globular domain of H1 is proposed to bind to the nucleosome as well as within the extra-core DNA
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
uracil DNA glycosylase activity on nucleosomal DNA depends on rotational orientation of targets
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
additional information
-
steady-state kinetics of wild-type and mutant enzymes
-
additional information
additional information
-
thermodynamics of UNG interactions with undamaged and damaged, e.g. 2'-deoxypseudouridine, substrates
-
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
metabolism
uracil DNA glycosylase initiates the uracil excision repair pathway
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
additional information
modeling of thymine in the active site pocket of Ung, overview
additional information
-
primary and secondary structure, and structural studies of ligand complexes, overview
additional information
-
structure comparison to the human and Herpes simplex virus enzymes, overview
additional information
-
the structural fold of UNG proteins is based on four beta-sheets sandwiched between two pairs of alpha-helices, the surface of the protein globule is traversed by a shallow and narrow positively charged groove, where substrate DNA binds. This groove harbors the enzyme's active site, which in turn contains a deep pocket that accommodates the uracil base, conformational change upon substrate binding, overall structure and conserved structural motifs, structure-function relationship, overview
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
strain B UDG free or in complex with uracil or glycerol, X-ray diffraction structure determination and analysis at 1.60, 1.50, and 1.43 A resolution, respectively
-
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
N123A
the mutation substantially dramatically reduces enzyme activity on A/U base pairs and other double-stranded uracil-containing base pairs
N123D/L191A
the mutation generates an enzyme that excises cytosine and distinguishes between cytosine and methylcytosine
Y66H
the mutant shows 170fold reduced uracil excision activity compared to the wild-type enzyme, but like the wild-type protein, it is susceptible to inhibition by uracil and AP-DNA
Y66W
the mutant shows 7fold reduced uracil excision activity compared to the wild-type enzyme, and lacks TDG activity. The Y66W protein is moderately compromised and attenuated in binding to AP-DNA. The Y66W mutant maintains strict specificity for uracil excision from DNA, but it is recalcitrant to inhibition by uracil and AP-DNA
D64N
-
site-directed mutagenesis, the mutant shows altered kinetics compared to the wild-type enzyme
H187Q
-
site-directed mutagenesis, the mutant shows altered kinetics compared to the wild-type enzyme
additional information
additional information
allelic exchange of ung with ung::kan, ungY66H:amp or ungY66W:amp alleles shows 5fold, 3.0fold, and 2.0fold, respectively, increase in mutation frequencies, widening of the substrate binding pocket can lead to aquirement of thymine DNA glycosylase activity, overview
additional information
-
UNG mutants with the set of bonds in the conserved 143GQ144 motif optimized for recognition of Cyt or Thy instead of Ura are able to excise normal pyrimidines from DNA and confer a spontaneous mutator phenotype to overexpressing Escherichia coli cells
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
HiTrap chelating column chromatography and HiTrap SP column chromatography
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
overexpression of wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
expression of wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
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REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Sobek, H.; Schmidt, M.; Frey, B.; Kaluza, K.
Heat-labile uracil-DNA glycosylase: purification and characterization
FEBS Lett.
388
1-4
1996
Escherichia coli
Pearl, J.H.
Structure and function in the uracil-DNA glycosylase superfamily
Mutat. Res.
460
165-181
2000
Escherichia coli, Homo sapiens, Human alphaherpesvirus 1 (P10186)
Porecha, R.H.; Stivers, J.T.
Uracil DNA glycosylase uses DNA hopping and short-range sliding to trap extrahelical uracils
Proc. Natl. Acad. Sci. USA
105
10791-10796
2008
Escherichia coli
Xiao, G.; Tordova, M.; Jagadeesh, J.; Drohat, A.C.; Stivers, J.T.; Gilliland, G.L.
Crystal structure of Escherichia coli uracil DNA glycosylase and its complexes with uracil and glycerol: structure and glycosylase mechanism revisited
Proteins
35
13-24
1999
Escherichia coli, Escherichia coli B / ATCC 11303
Huang, H.; Stivers, J.T.; Greenberg, M.M.
Competitive inhibition of uracil DNA glycosylase by a modified nucleotide whose triphosphate is a substrate for DNA polymerase
J. Am. Chem. Soc.
131
1344-1345
2009
Escherichia coli, Homo sapiens (P13051)
Cole, H.A.; Tabor-Godwin, J.M.; Hayes, J.J.
Uracil DNA glycosylase activity on nucleosomal DNA depends on rotational orientation of targets
J. Biol. Chem.
285
2876-2885
2010
Escherichia coli (P12295)
Zharkov, D.O.; Mechetin, G.V.; Nevinsky, G.A.
Uracil-DNA glycosylase: Structural, thermodynamic and kinetic aspects of lesion search and recognition
Mutat. Res.
685
11-20
2010
Escherichia coli, Homo sapiens
Bharti, S.K.; Varshney, U.
Analysis of the impact of a uracil DNA glycosylase attenuated in AP-DNA binding in maintenance of the genomic integrity in Escherichia coli
Nucleic Acids Res.
38
2291-2301
2010
Escherichia coli (P12295)
Xiang, Y.; Lu, Y.
Expanding targets of DNAzyme-based sensors through deactivation and activation of DNAzymes by single uracil removal: sensitive fluorescent assay of uracil-DNA glycosylase
Anal. Chem.
84
9981-9987
2012
Escherichia coli
Lee, D.H.; Liu, Y.; Lee, H.W.; Xia, B.; Brice, A.R.; Park, S.H.; Balduf, H.; Dominy, B.N.; Cao, W.
A structural determinant in the uracil DNA glycosylase superfamily for the removal of uracil from adenine/uracil base pairs
Nucleic Acids Res.
2
1081-1089
2015
Escherichia coli (P12295), Escherichia coli
Kimber, S.T.; Brown, T.; Fox, K.R.
A mutant of uracil DNA glycosylase that distinguishes between cytosine and 5-methylcytosine
PLoS ONE
9
e95394
2014
Escherichia coli (P12295)
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