Information on EC 3.2.2.28 - double-stranded uracil-DNA glycosylase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
3.2.2.28
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RECOMMENDED NAME
GeneOntology No.
double-stranded uracil-DNA glycosylase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
Specifically hydrolyses mismatched double-stranded DNA and polynucleotides, releasing free uracil
show the reaction diagram
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SYSTEMATIC NAME
IUBMB Comments
uracil-double-stranded DNA deoxyribohydrolase (uracil-releasing)
No activity on DNA containing a T/G mispair or single-stranded DNA containing either a site-specific uracil or 3,N4-ethenocytosine residue [2], significant role for double-stranded uracil-DNA glycosylase in mutation avoidance in non-dividing E. coli [3]. Uracil-DNA glycosylases are widespread enzymes that are found in all living organisms. Uracil-DNA glycosylase (EC 3.2.2.27) and 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 hide
59088-21-0
cf. EC 3.2.2.27
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain H37Ra, gene Rv1259 encoding UdgB
UniProt
Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
3,N4-ethenocytosine-containing single-stranded DNA + H2O
3,N4-ethenocytosine + single-stranded DNA with abasic site
show the reaction diagram
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the enzyme excised both 3,N4-ethenocytosine and uracil from DNA. 3,N4-ethenocytosine is significantly better as a substrate in terms of binding and hydrolysis. The tighter binding of the 3,N4-ethenocytosine containing substrate by MUG probably also accounts for its activity against single-stranded DNA containing 3,N4-ethenocytosine. Cleavage of the single-stranded substrate is 1500fold slower than the double-stranded substrate
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?
3,N4-ethenocytosine-mismatched double-stranded DNA + H2O
3,N4-ethenocytosine + double-stranded DNA with abasic site
show the reaction diagram
hypoxanthine-mismatched double-stranded DNA + H2O
hypoxanthine + double-stranded DNA with abasic site
show the reaction diagram
the enzyme also acts as a hypoxanthine DNA glycosylase with the strongest activity on the G/I base pair but no activity detected on the C/I base pair
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?
uracil-mismatched double-stranded DNA + H2O
uracil + double-stranded DNA with abasic site
show the reaction diagram
xanthine-mismatched double-stranded DNA + H2O
xanthine + double-stranded DNA with abasic site
show the reaction diagram
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?
xanthine-mismatched single-stranded DNA + H2O
xanthine + single-stranded DNA with abasic site
show the reaction diagram
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?
additional information
?
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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
hypoxanthine-mismatched double-stranded DNA + H2O
hypoxanthine + double-stranded DNA with abasic site
show the reaction diagram
Q5SJ65
the enzyme also acts as a hypoxanthine DNA glycosylase with the strongest activity on the G/I base pair but no activity detected on the C/I base pair
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?
uracil-mismatched double-stranded DNA + H2O
uracil + double-stranded DNA with abasic site
show the reaction diagram
Q5SJ65
the enzyme acts as a double-stranded uracil DNA glycosylase with a relatively low activity on the A/U base pair
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?
xanthine-mismatched double-stranded DNA + H2O
xanthine + double-stranded DNA with abasic site
show the reaction diagram
Q5SJ65
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?
xanthine-mismatched single-stranded DNA + H2O
xanthine + single-stranded DNA with abasic site
show the reaction diagram
Q5SJ65
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?
additional information
?
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Mug is expressed poorly in exponentially growing cells and has no apparent role in mutation avoidance in these cells. Mug is fairly abundant in stationary-phase cells and has an important anti-mutator role at this stage of cell growth
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INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
endonuclease IV
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endonuclease IV stimulates Dug activity by enhancing the rate and extent of uracil excision by promoting dissociation of Dug from the apyrimidinic-site/G-containing 34-mer. Catalytically active endonuclease IV is required to mediate Dug turnover
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0000025
3,N4-ethenocytosine-mismatched double-stranded DNA
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0.0000227
uracil-mismatched double-stranded DNA
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.016
3,N4-ethenocytosine-mismatched double-stranded DNA
Escherichia coli
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0.0028
uracil-mismatched double-stranded DNA
Escherichia coli
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pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
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poorly expressed in exponentially growing cells. Mug is overexpressed as cells enter stationary phase, and it is maintained at a fairly high level in resting cells. This is true of cells grown in rich or minimal media, and the principal regulation of mug is at the level of mRNA. Although the expression of mug is strongly dependent on the stationary-phase sigma factor, when cells are grown in minimal media, it shows only a modest dependence on stationary-phase sigma factor when cells are grown in rich media
Manually annotated by BRENDA team
PDB
SCOP
CATH
ORGANISM
UNIPROT
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
18670
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matrix-assisted laser desorption-ionization mass spectrometry
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
crystal structure of the Escherichia coli MUG enzyme complexed with an oligonucleotide containing a non-hydrolysable deoxyuridine analogue mismatched with guanine
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crystal structures of the mismatch-specific uracil DNA-glycosylase from Escherichia coli, and of a DNA complex, reveal a remarkable structural and functional homology to UDGs despite low sequence identity
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
60
purified UdgB, stable up to, MtuUdgB is thermo-tolerant
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
HiTrap chelating column chromatography
native and recombinant protein
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recombinant His6-tagged wild-type and mutant enzymes from Escherichia coli strains by nickel affinity chromatography
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli BH214 cells
gene Rv1259, DNA and amino aid sequence determination and analysis, cloning and overexpression of His6-tagged wild-type and mutant enzymes in Escherichia coli strains
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D75A
the mutant loses xanthine DNA glycosylase activity while retaining partial uracil and hypoxanthine DNA glycosylase activity
D75N
the mutant loses xanthine DNA glycosylase activity while retaining much of their uracil and hypoxanthine DNA glycosylase activity
D75Q
the mutant loses xanthine DNA glycosylase activity while retaining much of their uracil and hypoxanthine DNA glycosylase activity
N120A
the mutant loses xanthine DNA glycosylase activity while retaining partial uracil and hypoxanthine DNA glycosylase activity
additional information
construction of mutants by insertion of kanamycin resistance marker in the UdgB active site motif GQDPY, phenotypes, overview
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