Information on EC 3.6.1.25 - triphosphatase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
3.6.1.25
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RECOMMENDED NAME
GeneOntology No.
triphosphatase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
triphosphate + H2O = diphosphate + phosphate
show the reaction diagram
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phosphorous acid anhydride hydrolysis
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-
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SYSTEMATIC NAME
IUBMB Comments
triphosphate phosphohydrolase
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CAS REGISTRY NUMBER
COMMENTARY hide
62213-21-2
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
mold fungus
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Manually annotated by BRENDA team
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-
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Manually annotated by BRENDA team
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-
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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
ATP + H2O
?
show the reaction diagram
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low affinity
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-
?
triphosphate + H2O
diphosphate + phosphate
show the reaction diagram
tripolyphosphate + H2O
diphosphate + phosphate
show the reaction diagram
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very good substrate with Mg2+ as activator. The enzyme has a strong preference for linear tripolyphosphate compared with cyclic trimetaphosphate and to the linear tetraphosphate
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?
additional information
?
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the enzyme has low affinity for CTP, ATP or thiamine triphosphate. Nucleoside triphosphatase activity is negligible in the presence of 5 mM Mg2+, but a small activity is observed in the presence of 1 mM Mn2+, in particular with GTP. Guanosine 5-tetraphosphate and long chain polyphosphate (containing about 65 phosphate residues) are not hydrolyzed. The enzyme has no adenylyl cyclase activity
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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
triphosphate + H2O
diphosphate + phosphate
show the reaction diagram
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Co2+
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Co2+ is a better activator than Mn2+, although the maximum activity is less than 10% that measured in the presence of 5 mM Mg2+
Mg2+
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the enzyme hydrolyzes tripolyphosphate with high catalytic efficiency in the presence of 5 mM Mg2+
Mn2+
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very poor substituent for Mg2+ in the pH range 7.0-10, but there is a significant Mn2+-dependent activity at pH 10.0-10.5. Half-maximum activation is obtained at 0.4 mM
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Zn2+
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complete inhibition at 0.005 mM
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.8 - 1.2
ATP
0.02 - 74
tripolyphosphate
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.36 - 3.96
ATP
0.28 - 7900
tripolyphosphate
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.45 - 3.3
ATP
4
4 - 79000
tripolyphosphate
739
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.01
Cd2+
Nitrosomonas europaea
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His-tagged wild type enzyme, at pH 9.7 and 37C, in the presence of 5 mM Mg2+
0.02
Cu2+
Nitrosomonas europaea
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His-tagged wild type enzyme, at pH 9.7 and 37C, in the presence of 5 mM Mg2+
0.0015
Zn2+
Nitrosomonas europaea
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His-tagged wild type enzyme, at pH 9.7 and 37C, in the presence of 5 mM Mg2+
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.001
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R265S mutant, 2 mM methionine and 2 mM ATP as substrate
0.0012
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tripolyphosphatase activity of R265H mutant, 2 mM ATP as substrate
0.0019
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tripolyphosphatase activity of R265H mutant, 2 mM pyrophosphate as substrate
0.002
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R265H mutant, 2 mM methionine and 2 mM ATP as substrate
0.004
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tripolyphosphatase activity of R265H mutant, 2 mM metatripolyphosphate as substrate
0.029
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tripolyphosphatase activity of R265S mutant, 2 mM tripolyphosphate as substrate
0.067
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tripolyphosphatase activity of R265H mutant, 2 mM tripolyphosphate as substrate in the presence of 5 mM pyrophosphate
0.128
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tripolyphosphatase activity of R265H mutant, 2 mM tripolyphosphate as substrate
0.129
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tripolyphosphatase activity of R265H mutant, 2 mM tripolyphosphate as substrate in the presence of 5 mM ATP
0.156
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tripolyphosphatase activity of wildtype, 2 mM tripolyphosphate as substrate
0.588
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wildtype, 2 mM methionine and 2 mM ATP as substrate
additional information
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tripolyphosphate activity of R265H mutant is specific for tripolyphosphate and depends on magnesium ions, potassium is not required
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
9.7
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around pH 9.7
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Nitrosomonas europaea (strain ATCC 19718 / CIP 103999 / KCTC 2705 / NBRC 14298)
Nitrosomonas europaea (strain ATCC 19718 / CIP 103999 / KCTC 2705 / NBRC 14298)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
19000
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2 * 19000, SDS-PAGE
41600
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R265H mutant, gel filtration, column equilibrated with 2 mM tripolyphosphate
60000
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calculation from mRNA-sequence, subunit of D1 capping enzyme
90000
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heterodimer of R265H mutant and wildtype enzyme, size exclusion chromatography
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
heterodimer
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containing one R265H mutant subunit and one wildtype subunit, impaired AdoMet synthetase activity
homodimer
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2 * 19000, SDS-PAGE
monomer
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R265H mutant
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
hanging drop vapor diffusion method, selenomethionine-substituted protein, using 3.5 M sodium formate as a precipitation agent and 0.1 M Bis-Tris propane buffer, pH 7.0, at 20C
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
His-tagged R265H mutant purified with Ni2+ Sepharose column chromatography and wildtype enzyme purified from liver
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His-Trap column chromatography, gel filtration
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli BL21(DE3) cells
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His-tagged R265H mutant, expressed in E. coli BL21
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
K52R
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the His-tagged mutant loses 90-99% of its activity (catalytic efficiency is at least 1000times lower) compared to the wild type enzyme. Mn2+ does not induce a significant activation of this mutant
K85A
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the His-tagged mutant is about 10times less active than the wild type enzyme, but the optimal conditions for activity are essentially the same. The mutant is more strongly activated by Mn2+ than by Mg2+, and the inhibitory effects of Ca2+ and Zn2+ are less pronounced
K8A
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the His-tagged mutant remains highly active with tripolyphosphate as substrate and Mg2+ as activator, with a catalytic efficiency close to that of the recombinant wild type enzyme
R265H
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no AdoMet synthetase activity, normal tripolyphosphatase activity
R265S
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no AdoMet synthetase activity, reduced tripolyphosphatase activity