Information on EC 3.5.4.2 - adenine deaminase

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The expected taxonomic range for this enzyme is: Bacteria, Eukaryota

EC NUMBER
COMMENTARY hide
3.5.4.2
-
RECOMMENDED NAME
GeneOntology No.
adenine deaminase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
adenine + H2O = hypoxanthine + NH3
show the reaction diagram
-
-
-
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of amidines
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
adenine salvage
-
-
Metabolic pathways
-
-
purine metabolism
-
-
Purine metabolism
-
-
SYSTEMATIC NAME
IUBMB Comments
adenine aminohydrolase
-
CAS REGISTRY NUMBER
COMMENTARY hide
9027-68-3
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
Arthrobacter aurescens
-
-
-
Manually annotated by BRENDA team
-
SwissProt
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
no activity in other than some bacteria and some lower eukaryotes
-
-
-
Manually annotated by BRENDA team
strain A3
-
-
Manually annotated by BRENDA team
strain A3
-
-
Manually annotated by BRENDA team
this enzyme catalyses the reaction of EC 3.5.4.2. and EC 3.5.4.4; strain 23344c
SwissProt
Manually annotated by BRENDA team
strain BY4742, gene AAH1
-
-
Manually annotated by BRENDA team
strain J-350P
-
-
Manually annotated by BRENDA team
strain J-350P
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
-
deletion of the AAH locus in intact parasites establishes that AAH is not an essential gene and that -/-aah cells are capable of salvaging the same range of purine nucleobases and nucleosides as wild type Leishmania donovani. The aah null mutant is able to infect murine macrophages in vitro and in mice, although the parasite loads in both model systems are modestly reduced compared with wild type infections
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
2,6-diaminopurine + H2O
?
show the reaction diagram
2,6-diaminopurine + H2O
guanine + NH3
show the reaction diagram
2-amino-6-bromopurine + H2O
?
show the reaction diagram
2-amino-6-chloropurine + H2O
guanine + HCl
show the reaction diagram
6-amino-2-hydroxypurine + H2O
xanthine + NH3
show the reaction diagram
6-bromopurine + H2O
hypoxanthine + HBr
show the reaction diagram
6-chloropurine + H2O
?
show the reaction diagram
6-chloropurine + H2O
hypoxanthine + HCl
show the reaction diagram
6-iodopurine + H2O
hypoxanthine + HI
show the reaction diagram
6-methoxypurine + H2O
?
show the reaction diagram
6-[(E)-4-hydroxy-3-methylbut-2-enylamino]purine + H2O
?
show the reaction diagram
6-[(Z)-4-hydroxy-3-methylbut-2-enylamino]purine + H2O
?
show the reaction diagram
adenine + H2O
?
show the reaction diagram
adenine + H2O
hypoxanthine + NH3
show the reaction diagram
benzyladenine + H2O
hypoxanthine + 1-phenylmethaneamine
show the reaction diagram
kinetin + H2O
?
show the reaction diagram
kinetin + H2O
hypoxanthine + 1-furan-2-ylmethaneamine
show the reaction diagram
m-topolin + H2O
hypoxanthine + 2-(aminomethyl)phenol
show the reaction diagram
N-6-methyladenine + H2O
?
show the reaction diagram
N6-(2-isopentenyl)adenine + H2O
?
show the reaction diagram
N6-(2-isopentenyl)adenine + H2O
hypoxanthine + 3-methylbut-2-en-1-amine
show the reaction diagram
N6-isopentenyladenine + H2O
hypoxanthine + 3-methylbut-3-en-1-amine
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
6-chloropurine + H2O
?
show the reaction diagram
-
-
-
-
-
adenine + H2O
?
show the reaction diagram
adenine + H2O
hypoxanthine + NH3
show the reaction diagram
kinetin + H2O
?
show the reaction diagram
N6-(2-isopentenyl)adenine + H2O
?
show the reaction diagram
Q9P6I7
partial conversion, at endo-side of N6 amino groups, release of N6-side chain amines
-
-
?
N6-(2-isopentenyl)adenine + H2O
hypoxanthine + 3-methylbut-2-en-1-amine
show the reaction diagram
P53909
partial conversion, at endo-side of N6 amino groups, release of N6-side chain amines
-
-
?
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
-
no dialyzable cofactors required
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Co2+
-
11% of the activity with Mn2+
Fe3+
-
[FeII/FeII]-ADE is oxidized to [FeIII/FeIII]-ADE with ferricyanide with inactivation of the deaminase activity
Zn2+
-
required for activity
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1,10-phenanthroline
2'-deoxycoformycin
-
-
3-methyl-1-phenyl-2-(phenylseleno)oct-2-en-1-one
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an inhibition of ADA activity using 100 nM is observed, an increase in cellular viability is shown when 0.03 mM is used
4-Aminopyrazolo[3,4-d]pyrimidine
-
-
6-Chloropurine
-
-
6-Dimethylaminopurine
-
-
6-Methylaminopurine
-
-
coformycin
Cu2+
64% residual activity at 1 mM
Deoxycoformycin
Fe2+
7% residual activity at 1 mM
hypoxanthine
-
competitive, Ki 4 mM
Mercuric acetate
-
-
Mercuric chloride
-
-
Monoiodoacetic acid
-
-
Ni2+
48% residual activity at 1 mM
p-chloromercuribenzoate
-
-
purine
additional information
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.07 - 1.1
2,6-Diaminopurine
1
2-amino-6-chloropurine
-
-
1.5
2-hydroxy-6-aminopurine
-
-
0.5
6-bromopurine
-
-
0.026 - 1
6-Chloropurine
0.13 - 0.27
6-methoxypurine
0.0154 - 2.2
adenine
0.088 - 0.09
N-6-methyladenine
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.8 - 17
2,6-Diaminopurine
0.42 - 1.9
6-Chloropurine
0.52 - 7.1
6-methoxypurine
0.022 - 200
adenine
0.48 - 1.28
N-6-methyladenine
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
19 - 120
2,6-Diaminopurine
7.3 - 16
6-Chloropurine
4 - 26
6-methoxypurine
0.026 - 3100
adenine
1.3 - 15
N-6-methyladenine
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0231
2'-deoxycoformycin
-
pH 7.0, 25C
0.00013
6-Chloropurine
-
pH 7.5, 30C
4
hypoxanthine
-
competitive
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.014 - 0.14
-
-
0.071
-
in crude extracts
1.88
after 5.26fold purification, at pH 7.4 and 40C
5.97
-
pH 7.5, 37C
8.34
recombinant protein, adenine as substrate, 0.067 mM in 0.2 M phosphate buffer, 33C, pH 7.0, adenine most favoured substrate, also hydrolyzes N6-substituted adenines from the cytokinin group of plant hormones, reaction products identified by mass spectroscopy, relative reaction rates given
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.5 - 7
6.5 - 7.5
-
substrate: 2-amino-6-chloropurine
6.7
0.2 M phosphate buffer, 33C; assay at
7 - 7.5
-
medium at
7
-
enzyme not purified
7.5
-
substrate: 6-chloropurine, 6-iodopurine, 2,6-diaminopurine
7.6
-
assay at
9
-
Tris-HCl buffer at
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 8.5
-
pH 5: about 30% of maximal activity, pH 8.5: about 20% of maximal activity
6 - 8
-
potassium phosphate buffer, maximal activity pH 7
7 - 9
-
Tris-HCl buffer, maximal activity pH 9
7.5 - 9
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glycine-NaOH buffer, maximal activity pH 7
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 37
0.2 M phosphate buffer, pH 7.0
33
0.2 M phosphate buffer, pH 6.7; assay at
35
ranges from 30-37C
37
-
assay at
40
-
enzyme not purified
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 37
determined at pH 7.0
PDB
SCOP
CATH
ORGANISM
UNIPROT
Agrobacterium fabrum (strain C58 / ATCC 33970)
Agrobacterium fabrum (strain C58 / ATCC 33970)
Agrobacterium fabrum (strain C58 / ATCC 33970)
Enterococcus faecalis (strain ATCC 700802 / V583)
Pseudomonas aeruginosa (strain ATCC 15692 / PAO1 / 1C / PRS 101 / LMG 12228)
Pseudomonas aeruginosa (strain ATCC 15692 / PAO1 / 1C / PRS 101 / LMG 12228)
Pseudomonas aeruginosa (strain ATCC 15692 / PAO1 / 1C / PRS 101 / LMG 12228)
Pseudomonas aeruginosa (strain ATCC 15692 / PAO1 / 1C / PRS 101 / LMG 12228)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37000
-
gel filtration
39630
calculated; deduced from sequence
40800
-
SDS-PAGE
41200
calculated; deduced from sequence
120000
-
gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homodimer
-
2 * 60000, SDS-PAGE
monomer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
X-ray structures of Pa0148 (Pseudomonas aeruginos) and AAur1117 (Arthrobacter aurescens) are determined and reveal nearly identical distorted (beta/alpha)8 barrels with a single zinc ion that is characteristic of members of the amidohydrolase superfamily
Arthrobacter aurescens
-
modeling of structure reveals (beta/alpha)8 barrel and amino acids H15, H17, H214, D295, E217 and H238 involved in catalytic mechanism
the three-dimensional structure of adenine deaminase from Agrobacterium tumefaciens (Atu4426) is determined by X-ray crystallography at 2.2 A resolution
-
X-ray structures of Pa0148 (Pseudomonas aeruginos) and AAur1117 (Arthrobacter aurescens) are determined and reveal nearly identical distorted (beta/alpha)8 barrels with a single zinc ion that is characteristic of members of the amidohydrolase superfamily
-
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.5 - 7.5
the enzyme has maximal stability at 30C for 24 h in HEPES buffer at pHs between 5.5 and 7.5
734047
additional information
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 50
the enzyme loses 35% of its catalytic function in 1 h at 40C, whereas at 30C, the activity of the enzyme remains stable. At 50C, the activity drops within 10 min to zero
37
-
pH 7.0, potassium phosphate buffer, unstable
65
-
20% loss of activity after 5 min
additional information
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
after dissolving the freeze-dried enzyme a significant decrease od activity is observed within 3-4 h
bovine serum albumin protects against heat inactivation in dilute solutions
-
dialysis, 24 h against 0.9% KCl, stable
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-15C, ammonium sulfate, for at least 2 months
-
4C, 10 mM phoshate buffer, pH7.5, 20 mM 2-mercaptoethanol, 1-2 weeks, minimal losses
activity of purified protein is stable for 1-2 weeks when stored at 4C at concentrations above 10 mg/ml with the addition of 20% (v/v) glycerol. Prolonged storage in solution is possible at -20C with 50% (v/v) glycerol and at -80C with 10-20% (v/v) glycerol or as freeze-dried powder
at 4C in potassium buffer above a concentration of 10 mg/ml and in presence of 20% glycerol stable for 1-2 weeks, prolonged storage in solution at -20C in presence of 50% glycerol and at -80C in presence of 10-20% glycerol, or as freeze-dried powder
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate precipitation, DEAE-Sepharose column chromatography, and Superdex 200 gel filtration
gel filtration, SDS-PAGE; using Ni2+-nitrilotriacetate (Ni2+-NTA) column chromatography
His-tagged, recombinant protein
recombonant protein
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli Top10 cells
expressed in Escherichia coli, pDrive, pet100/D-TOPO, pet151/D-TOPO vectors; overexpression in Escherichia coli BL21 (DE3)
expression in Escherichia coli
-
overexpression in Escherichia coli
-
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
Western blot analysis establishes that AAH is expressed in both life cycle stages of Leishmania donovani
-
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D118N
-
mutant is able to bind 2 metals per active site, kcat (adenine): 173/sec
D284A
-
mutants is able to bind two equivalents of Mn2+ or Fe2+ in the active site but mutant is unable to catalyze the deaminase reaction
D285A
-
mutant is able to bind 2 metals per active site, kcat (adenine): 37/sec
D474N
-
mutant is able to bind 2 metals per active site, kcat (adenine): 171/sec
E121Q
-
mutant is able to bind 2 metals per active site, kcat (adenine): 57/sec
E185Q
-
mutant shows total loss of catalytic activity as well as the ability to bind divalent metals in the active site
E236Q
-
mutants is able to bind two equivalents of Mn2+ or Fe2+ in the active site but mutant is unable to catalyze the deaminase reaction
H120N
-
mutant is able to bind 2 metals per active site, kcat (adenine): 0.13/sec
H214N
-
mutant shows total loss of catalytic activity as well as the ability to bind divalent metals in the active site
H214Q
-
mutant shows total loss of catalytic activity as well as the ability to bind divalent metals in the active site
H235C
-
mutant shows total loss of catalytic activity as well as the ability to bind divalent metals in the active site
H235D
-
mutant shows total loss of catalytic activity as well as the ability to bind divalent metals in the active site
H235N
-
mutant shows total loss of catalytic activity as well as the ability to bind divalent metals in the active site
H235Q
-
mutant shows total loss of catalytic activity as well as the ability to bind divalent metals in the active site
H473N
-
mutant is able to bind 2 metals per active site, kcat (adenine): 178/sec
H90C
-
mutant is unable to bind either iron or manganese
H90D
-
mutant is unable to bind either iron or manganese
H90N
-
mutant is able to bind two equivalents of Mn2+ or Fe2+ per monomer and shows a kcat of about 5% of the wild-type enzyme
H90Q
-
mutant is unable to bind either iron or manganese
H92C
-
mutant shows total loss of catalytic activity as well as the ability to bind divalent metals in the active site
H92D
-
mutant shows total loss of catalytic activity as well as the ability to bind divalent metals in the active site
H92N
-
mutant shows total loss of catalytic activity as well as the ability to bind divalent metals in the active site
H92Q
-
mutant shows total loss of catalytic activity as well as the ability to bind divalent metals in the active site
S95A
-
mutant is able to bind 2 metals per active site, kcat (adenine): 78/sec
additional information
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