Information on EC 2.7.1.127 - inositol-trisphosphate 3-kinase and Organism(s) Rattus norvegicus and UniProt Accession P17105

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Rattus norvegicus
UNIPROT: P17105


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The enzyme appears in selected viruses and cellular organisms

EC NUMBER
COMMENTARY hide
2.7.1.127
-
RECOMMENDED NAME
GeneOntology No.
inositol-trisphosphate 3-kinase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + 1D-myo-inositol 1,4,5-trisphosphate = ADP + 1D-myo-inositol 1,3,4,5-tetrakisphosphate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Phosphorylation
-
phospho-group transfer
-
phospho group transfer
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
1D-myo-inositol hexakisphosphate biosynthesis I (from Ins(1,4,5)P3)
-
-
1D-myo-inositol hexakisphosphate biosynthesis II (mammalian)
-
-
D-myo-inositol (1,3,4)-trisphosphate biosynthesis
-
-
Inositol phosphate metabolism
-
-
Metabolic pathways
-
-
SYSTEMATIC NAME
IUBMB Comments
ATP:1D-myo-inositol-1,4,5-trisphosphate 3-phosphotransferase
Activated by Ca2+. Three isoforms have been shown to exist [3].
CAS REGISTRY NUMBER
COMMENTARY hide
106283-10-7
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
-
inositol(1,4,5)trisphosphate 3-kinases (Itpks) occur in three isoenzyme forms, Itpka/b and c, in human, rat and mouse. They share a catalytic domain relatively well conserved at the C-terminal end and a quite isoenzyme specific regulatory domain at the N-terminal end of the protein
malfunction
-
neurite length is significantly decreased in cells overexpressing isozymes Itpka and Itpkb but not Itpkc or IPMK. This result does not depend on the overexpression level of any of the kinases. PC12 cells overexpressing GFP-tagged kinase-dead mutants Itpka/b have shorter neurites than GFP control cells
metabolism
-
1D-myo-inositol 1,3,4,5-tetrakisphosphate, Ins(1,3,4,5)P4 can interact with a relatively specific Ins(1,3,4,5)P4 binding protein Rasa3, alternatively, Ins(1,3,4,5)P4 can also compete with phosphoinositides to the binding of PH domain containing proteins such as Akt, protein kinase B. In neutrophils and hematopoietic progenitors, elevated levels of Ins(1,3,4,5)P4 inhibit the recruitment of Akt at the plasma membrane, and its activation, acting as a competitor of PtdIns(3,4,5)P3 binding to its PH domain
physiological function
evolution
-
inositol(1,4,5)trisphosphate 3-kinases (Itpks) occur in three isoenzyme forms, Itpka/b and c, in human, rat and mouse. They share a catalytic domain relatively well conserved at the C-terminal end and a quite isoenzyme specific regulatory domain at the N-terminal end of the protein
malfunction
-
neurite length is significantly decreased in cells overexpressing isozymes Itpka and Itpkb but not Itpkc or IPMK. This result does not depend on the overexpression level of any of the kinases. PC-12 cells overexpressing GFP-tagged kinase-dead mutants Itpka/b have shorter neurites than GFP control cells
metabolism
-
1D-myo-inositol 1,3,4,5-tetrakisphosphate, Ins(1,3,4,5)P4 can interact with a relatively specific Ins(1,3,4,5)P4 binding protein Rasa3, alternatively, Ins(1,3,4,5)P4 can also compete with phosphoinositides to the binding of PH domain containing proteins such as Akt, protein kinase B. In neutrophils and hematopoietic progenitors, elevated levels of Ins(1,3,4,5)P4 inhibit the recruitment of Akt at the plasma membrane, and its activation, acting as a competitor of PtdIns(3,4,5)P3 binding to its PH domain
physiological function
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ATP + 1D-myo-inositol 1,4,5-trisphosphate
ADP + 1D-myo-inositol 1,3,4,5-tetrakisphosphate
show the reaction diagram
ATP + 1D-myo-inositol 1,4,5-triphosphate
ADP + 1D-myo-inositol 1,3,4,5-tetrakisphosphate
show the reaction diagram
-
-
-
r
ATP + 1D-myo-inositol 1,4,5-trisphosphate
ADP + 1D-myo-inositol 1,3,4,5-tetrakisphosphate
show the reaction diagram
ATP + 1D-myo-inositol 2,4,5-trisphosphate
ADP + 1D-myo-inositol 2,4,5,6-tetrakisphosphate
show the reaction diagram
-
recombinant, catalytically active fragment of isoform C, in the presence of Ins(1,3,4,5)P4, authentic side activity of isoform C
-
?
ATP + D-2-deoxy-myo-inositol 1,4,5-trisphosphate
ADP + D-2-deoxy-myo-inositol 1,3,4,5-tetrakisphosphate
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
ATP + 1D-myo-inositol 1,4,5-trisphosphate
ADP + 1D-myo-inositol 1,3,4,5-tetrakisphosphate
show the reaction diagram
ATP + 1D-myo-inositol 1,4,5-triphosphate
ADP + 1D-myo-inositol 1,3,4,5-tetrakisphosphate
show the reaction diagram
Q80ZG2
-
-
-
r
ATP + 1D-myo-inositol 1,4,5-trisphosphate
ADP + 1D-myo-inositol 1,3,4,5-tetrakisphosphate
show the reaction diagram
additional information
?
-
-
the enzyme might be involved in brain development, memory, and learning
-
-
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Calmodulin
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
-
assay in presence of Triton X-100 and ethylene glycol bis-(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
(2'S)-1D-1,2-O-[(2'-phosphoryloxy)propane-1',3'-diyl]-myo-inositol 4,5-bisphosphate
-
synthetic bicyclic inositol trisphosphate S epimer, IC50 is 0.156 mM
1D-myo-inositol 1,3,4,5-tetrakisphosphate
1D-myo-inositol 1,4,5-trisphosphate
-
recombinant, catalytically active fragment of isoform C, substrate inhibition by high concentrations
3',4',7,8-tetrahydroxyflavone
aurintricarboxylic acid
chlorogenic acid
-
D-2-deoxyinositol 1,3,4,5-tetrakisphosphate
-
strong inhibition of isozyme A, IC50 is 0.0054 mM
D-2-deoxyinositol 1,4,5-trisphosphate
-
strong inhibition of isozyme A, IC50 is 0.0017 mM
D-3-deoxyinositol 1,4,6-trisphosphate
-
strong inhibition of isozyme A, IC50 is 0.0014 mM
D-6-deoxyinositol 1,3,4,5-tetrakisphosphate
-
strong inhibition of isozyme A, IC50 is 0.0051 mM
D-myo-inositol 1,4,5-trisphosphate
-
strong inhibition, purified recombinant enzyme, IC50 value is 0.003 mM in absence of Ca2+
D-myo-inositol 2,4,5-trisphosphate
-
IC50 is 0.117 mM
D-scyllo-inositol 1,2,3,4-tetrakisphosphate
-
purified recombinant enzyme, IC50 value is above 0.1 mM in absence of Ca2+
D-scyllo-inositol 1,2,4-trisphosphate
-
purified recombinant enzyme, IC50 value is 0.044 mM in absence of Ca2+
EGTA
-
Ca2+/calmodulin-activated enzyme
epicatechin-3-gallate
epigallocatechin-3-gallate
gossypol
hypericin
inositol phosphate
-
inhibitory effects of all possible 38 regioisomers of synthetic inositol phosphates, only inositol trisphosphates and tetrakisphosphates inhibit, not or very weak: inositol monophosphates, bisphosphates and pentakisphosphates, the recognition of myo-inositol phosphate regioisomers is highly structure-selective
KN-62
-
calmodulin-dependent protein kinase II inhibitor; prevents the carbachol- or UTP-mediated activation
KN-93
-
calmodulin-dependent protein kinase II inhibitor; prevents the carbachol- or UTP-mediated activation
L-scyllo-inositol 1,2,3,4-tetrakisphosphate
-
purified recombinant enzyme, IC50 value is above 0.1 mM in absence of Ca2+
L-scyllo-inositol 1,2,4-trisphosphate
-
purified recombinant enzyme, IC50 value is 0.006 mM in absence of Ca2+
myricetin
N-(6-Aminohexyl)-5-chloro-1-naphthalenesulfonamide
-
i.e. W-7, calmodulin-antagonist
protein kinase A
-
isoforms A and B are differentially regulated via phosphorylation by the cAMP-dependent protein kinase, isoform A: stimulation in the presence or absence of Ca2+/calmodulin, isoform B: no effect on activity in the absence of Ca2+/calmodulin, 45% inhibition in the presence of Ca2+/calmodulin
-
Protein kinase C
-
quercetin
scyllo-inositol 1,2,3,5-tetrakisphosphate
-
weak inhibition, purified recombinant enzyme, IC50 value is 0.028 mM in absence of Ca2+
scyllo-inositol 1,2,3-trisphosphate
-
purified recombinant enzyme, IC50 value is 0.027 mM in absence of Ca2+
scyllo-inositol 1,2,4,5-tetrakisphosphate
-
weak inhibition, purified recombinant enzyme, IC50 value is 0.039 mM in absence of Ca2+
scyllo-inositol 1,3,5-trisphosphate
-
purified recombinant enzyme, IC50 value is 0.090 mM in absence of Ca2+
TSH
-
thyroid-stimulating hormone, inhibits at a physiological concentration, inhibition is mimicked by dibuturyl cyclic AMP and forskolin, mechanism
additional information
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Calmodulin
-
the enzyme is activated by Ca2+/calmodulin. The isozyme contains a CaM binding domain
Carbachol
-
activates the enzyme up to 7fold in rat brain cortical slices
1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane
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5-HT2 agonist DOI, 3fold activation
12-O-tetradecanoyl phorbol 13-acetate
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stimulating in presence of cAMP
12-O-tetradecanoylphorbol-13-acetate
-
2fold activation, isoform B
1D-myo-inositol 1,3,4,5-tetrakisphosphate
-
recombinant, catalytically active fragment of isoform C, allosteric product activation in the absence of Ca2+/calmodulin, which per se activates enzyme and abolishes the allosteric effect
5-HT
-
2-3fold activation
Ca2+/CaM-dependent kinase II
-
i.e. CaMKII, activates the enzyme by phosphorylation of threonine residue
-
Calmodulin
CaM kinase II
-
cAMP-dependent protein kinase
-
i.e. PKA, activates the enzyme by phosphorylation at Ser109, simultaneous phosphorylation at SEr109 and Ser175 inactivates the enzyme
-
Carbachol
protein kinase A
-
isoforms A and B are differentially regulated via phosphorylation by the cAMP-dependent protein kinase, isoform A: phosphorylated to the extent of 0.9 mol/mol, 2.5fold stimulation in the absence of Ca2+/calmodulin, 1.5fold stimulation in the presence of Ca2+/calmodulin, isoform B: phosphorylated to the extent of 1 mol/mol, no effect on activity in the absence of Ca2+/calmodulin, 45% inhibition in the presence of Ca2+/calmodulin
-
Protein kinase C
-
phosphorylation of isoenzyme B by calmodulin kinase II and protein kinase C added together results in a maximal 60-70fold activation, but protein kinase C alone inhibits in the presence of Ca2+ and calmodulin, no effect on the sensitivity to the Ca2+/calmodulin complex
-
serotonin
-
potentiates enzyme activity mediated through the activation of 5-HT2 receptors, enzyme up-regulation occurs through activation of PLC-coupled serotoninergic receptors and requires the phosphorylation of the enzyme by the ubiquitous multimeric protein kinase CaMKII
additional information
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0002 - 0.011
1D-myo-inositol 1,4,5-trisphosphate
0.0015 - 0.005
1D-myo-inositol 2,4,5-trisphosphate
0.033 - 0.0876
ATP
0.0002 - 0.0017
InsP3
additional information
additional information
All potent inhibitors increases the KM-value for ATP
-
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.12
N-(6-Aminohexyl)-5-chloro-1-naphthalenesulfonamide
-
-
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.156
(2'S)-1D-1,2-O-[(2'-phosphoryloxy)propane-1',3'-diyl]-myo-inositol 4,5-bisphosphate
Rattus norvegicus
-
synthetic bicyclic inositol trisphosphate S epimer, IC50 is 0.156 mM
0.013
1D-myo-inositol 1,3,4,5-tetrakisphosphate
Rattus norvegicus
-
product inhibition, IC50 is 0.013 mM
0.00019 - 0.00061
3',4',7,8-tetrahydroxyflavone
0.000062
aurintricarboxylic acid
Rattus norvegicus
Q80ZG2
IP3K-C, maximal inhibition 100%
0.0054
D-2-deoxyinositol 1,3,4,5-tetrakisphosphate
Rattus norvegicus
-
strong inhibition of isozyme A, IC50 is 0.0054 mM
0.0017
D-2-deoxyinositol 1,4,5-trisphosphate
Rattus norvegicus
-
strong inhibition of isozyme A, IC50 is 0.0017 mM
0.0014
D-3-deoxyinositol 1,4,6-trisphosphate
Rattus norvegicus
-
strong inhibition of isozyme A, IC50 is 0.0014 mM
0.0051
D-6-deoxyinositol 1,3,4,5-tetrakisphosphate
Rattus norvegicus
-
strong inhibition of isozyme A, IC50 is 0.0051 mM
0.003
D-myo-inositol 1,4,5-trisphosphate
Rattus norvegicus
-
strong inhibition, purified recombinant enzyme, IC50 value is 0.003 mM in absence of Ca2+
0.117
D-myo-inositol 2,4,5-trisphosphate
Rattus norvegicus
-
IC50 is 0.117 mM
0.1
D-scyllo-inositol 1,2,3,4-tetrakisphosphate
Rattus norvegicus
-
purified recombinant enzyme, IC50 value is above 0.1 mM in absence of Ca2+
0.044
D-scyllo-inositol 1,2,4-trisphosphate
Rattus norvegicus
-
purified recombinant enzyme, IC50 value is 0.044 mM in absence of Ca2+
0.00069
ellagic acid
Rattus norvegicus
Q80ZG2
IP3K-C, maximal inhibition 85%
0.000385
epicatechin-3-gallate
Rattus norvegicus
Q80ZG2
IP3K-C, maximal inhibition 100%
0.00021
epigallocatechin-3-gallate
Rattus norvegicus
Q80ZG2
IP3K-C, maximal inhibition 100%
0.000175
gossypol
Rattus norvegicus
Q80ZG2
IP3K-C, maximal inhibition 100%
0.00017
hypericin
Rattus norvegicus
Q80ZG2
IP3K-C, maximal inhibition 100%
0.1
L-scyllo-inositol 1,2,3,4-tetrakisphosphate
Rattus norvegicus
-
purified recombinant enzyme, IC50 value is above 0.1 mM in absence of Ca2+
0.006
L-scyllo-inositol 1,2,4-trisphosphate
Rattus norvegicus
-
purified recombinant enzyme, IC50 value is 0.006 mM in absence of Ca2+
0.00029
myricetin
Rattus norvegicus
Q80ZG2
IP3K-C, maximal inhibition 62%
0.00039
quercetin
Rattus norvegicus
Q80ZG2
IP3K-C, maximal inhibition 71%
0.00017
Rose bengal
Rattus norvegicus
Q80ZG2
IP3K-C, maximal inhibition 100%
0.028
scyllo-inositol 1,2,3,5-tetrakisphosphate
Rattus norvegicus
-
weak inhibition, purified recombinant enzyme, IC50 value is 0.028 mM in absence of Ca2+
0.027
scyllo-inositol 1,2,3-trisphosphate
Rattus norvegicus
-
purified recombinant enzyme, IC50 value is 0.027 mM in absence of Ca2+
0.039
scyllo-inositol 1,2,4,5-tetrakisphosphate
Rattus norvegicus
-
weak inhibition, purified recombinant enzyme, IC50 value is 0.039 mM in absence of Ca2+
0.09
scyllo-inositol 1,3,5-trisphosphate
Rattus norvegicus
-
purified recombinant enzyme, IC50 value is 0.090 mM in absence of Ca2+
additional information
chlorogenic acid
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.00072
-
in absence of calmodulin
0.0008
-
liver
0.002
-
brain cortex
1.4
-
isoform B, in absence of the Ca2+/calmodulin complex
1.7
-
pH 7.4, 37°C, purified recombinant isoenzyme A overexpressed in CHO cells
3.5
-
30°C, fully activated enzyme in presence of Ca2+/calmodulin
3.68
-
pH 7.5, 37°C
15.77
-
pH 7.5, 37°C, recombinant isoform B
20
-
recombinant IP3K overexpressed in Escherichia coli, in absence of calmodulin
28.9
-
pH 7.5, 37°C, in absence of calmodulin
36.32
-
pH 7.5, 37°C, recombinant isoform A
60
-
pH 7.5, 37°C, in presence of calmodulin
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.4
-
assay at
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.5 - 8
-
inactive at pH 6.5 and below, above pH 6.5 the activity rises steeply to a broad optimum at pH 8
7 - 9
-
about half-maximal activity at pH 7 and about 60% of maximal activity at pH 9
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
-
Itpka is particularly active in neurons of the hippocampus and cerebellum
Manually annotated by BRENDA team
-
dendritic spines
Manually annotated by BRENDA team
-
high expression level
Manually annotated by BRENDA team
-
isozyme IP33K-C
Manually annotated by BRENDA team
-
thyroid cells
Manually annotated by BRENDA team
-
endogenous isoenzyme A is localized to the dentritic spines of pyramidal neurons in primary hippocampal cultures from neonatal rats, highest expression of recombinant isoenzyme A in astrocytes
Manually annotated by BRENDA team
-
isoform C, low expression
Manually annotated by BRENDA team
-
insulin-secreting
Manually annotated by BRENDA team
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
-
cytoskeletal localization for isozyme Itpka
Manually annotated by BRENDA team
isoform inositol 1,4,5-trisphosphate 3-kinase A is a microtubule-associated protein, and the N terminus of inositol 1,4,5-trisphosphate 3-kinase A is a critical region for direct binding to tubulin in dendritic shaft of hippocampal neurons. Protein kinase A phosphorylates residue Ser119 within inositol 1,4,5-trisphosphate 3-kinase A, leading to a significant reduction of microtubule binding affinity
Manually annotated by BRENDA team
-
recombinant isoenzyme A has an N-terminal 66-amino acid F-actin-binding region, isoform A co-localizes with F-actin
Manually annotated by BRENDA team
-
recombinant isoenzyme A has an N-terminal 66-amino acid F-actin-binding region that localizes the kinase to dentritic spines, endogenous isoenzyme A is localized to the dentritic spines of pyramidal neurons in primary hippocampal cultures
Manually annotated by BRENDA team
-
not activated isoenzyme B: 65% soluble, 35% particulate fraction, carbachol-activated isoenzyme B shows a redistribution of enzyme from soluble to particulate fraction, only 10% remain soluble
-
Manually annotated by BRENDA team
-
recombinant isoenzyme A associates with
Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
44000
-
x * 44000, SDS-PAGE
50900
-
x * 50900, isozyme IP33K-A, x * 74000, isozyme IP33K-B, x * 74500, isozyme IP33K-C
59000
-
x * 59000, recombinant isoform A, x * 92000, recombinant isoform B, SDS-PAGE
70000
-
gel filtration
74000
-
x * 50900, isozyme IP33K-A, x * 74000, isozyme IP33K-B, x * 74500, isozyme IP33K-C
88000
-
x * 88000, isoform B, SDS-PAGE, Western blot analysis
92000
-
x * 59000, recombinant isoform A, x * 92000, recombinant isoform B, SDS-PAGE
100000
-
x * 100000, SDS-PAGE
150000 - 160000
-
in the presence of Ca2+/calmodulin, gel filtration, the interaction of calmodulin with the monomeric catalytic subunit promotes dimerization of the enzyme
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
-
x * 50000, isozyme Itpka from rat brain, SDS-PAGE
monomer
-
1 * 53000, SDS-PAGE
additional information
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phosphoprotein
proteolytic modification
-
Itpka undergoes proteolysis in intact cells, leading to the accumulation of a stable C-terminal domain of 83 kDa to the endoplasmic reticulum, incubation of Itpka at 37°C for 30 min totally destroys Ins(1,4,5)P3 3-kinase activity unless the enzyme is mixed with 0.1% SDS
phosphoprotein
proteolytic modification
-
Itpkb is very sensitive to proteolysis
additional information
the enzyme is cleaved at the Asp-Pro clusters, residues 610, 617, and 765, at low pH and different temperatures producing 7 enzyme fragments of 17.4-82.3 kDa of the full-length enzyme of about 100 kDa, overview
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified recombinant unlabeled or selennomethionine-labeled mutant catalytic domain residues 185-459, 10 mg/ml protein in 25 mM Tris-HCl, pH 8.0, 150 mM NaCl, and 10 mM DTT, crystallization solution contains 2.2 M ammonium sulfate, 0.2 M sodium formate, 10 mM DTT, 10 mM ADP, 10 mM 1D-myo-inositol 1,3,4,5-tetrakisphosphate, and 10 mM MgCl2, cryoprotection of crystals by successive transfer into 5-15% glycerol in mother liquor, X-ray diffraction structure determination and analysis at 2.2 A resolution
-
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
0.1% CHAPS or 0.1% Triton X-100 stabilizes, 0.5 mg/ml bovine serum albumin is less efficient in stabilizing
-
2-mercaptoethanol and Triton X-100 or CHAPS stabilize during purification
-
bovine serum albumin stabilizes dilute enzyme solutions
-
calpain inhibitors stabilize during purification
-
ethylene glycol, 10%, KCl or sucrose, 0.25 M, does not stabilize
-
isoform A is more susceptible to proteolysis during purification than isoform B
-
stable to dialysis against 0.15 M sucrose
-
ORGANIC SOLVENT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Triton X-100
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-15°C, partially purified brain enzyme, very stable
-
-20°C or 4°C, 0.1% CHAPS or Triton X-100, 4 days, less than 5% loss of activity
-
-20°C, 4 days, 58% loss of activity
-
-70°C, buffer containing detergent and reducing agent, pH 7-8, at least 3 months, stable
-
4°C, 20 mM Tris-HCl buffer, pH 7.2, 1 mM Mg(CH3COO)2, 1 mM DTT, 0.1 mM EGTA, 0.5 mg/ml bovine serum albumin, 3 months, less than 20% loss of activity
-
4°C, 4 days, 20% loss of activity
-
4°C, 48 h, 10-20% loss of activity
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
elution of recombinant Itpka isozyme with 0.5% SDS and 0.1 mM EGTA from calmodulin affinity column
-
20fold; partial
-
2700fold; partial
-
403fold; purified in the presence of SDS as a doublet of a MW of approximately 50000
-
83fold; recombinant IP3K overexpressed in Escherichia coli
-
By phosphocellulose.
isoenzyme B
-
native enzyme partially by nuclear fraction preparation
-
recombinant GST-fusion isoform B comprising residues 108-170 by glutathione affinity chromatography, recombinant truncated Strep-tagged isozyme B by affinity chromatography
recombinant IP3K overexpressed in Escherichia coli
-
recombinant isoforms A and B, overexpressed in B31 rat fibroblast, isoform A: 11005fold, isoform B: 43806fold
-
recombinant N-terminally His-tagged and S-tagged C-terminal catalytic domain 3KA-cat30, comprising residues Ser185-Arg459, from Escherichia coli strain BL21(DE3) by nickel and S-protein affinity chromatography, and gel filtration
-
unlabeled or selenomethionine-labeled recombinant residues 185-459 of the catalytic domain of isozyme A fused to GST from Escherichia coli by glutathione affinity chromatography, removal of the GST-tag by recombinant TEV protease, further purification by a second step of glutathione affinity chromatography, adsorption chromatography, and gel filtration
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
gene Itpka
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generation of knock-out rats and point mutant, IP3K-A is overexpressed in DIV 22 hippocampal neurons by infection with adenovirus containing full-length IP3K-A
a cDNA encoding InsP3 3-kinase B is cloned from a thymus cDNA library and expressed in Escherichia coli, sequence of the 673 amino acids protein
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cDNAs encoding enzyme isoforms A from brain and B are cloned and overexpressed in B31 cells, a rat-1 fibroblast cell line expressing pp60v-src
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cloning of a full-length cDNA encoding IP3K isoform C from tongue epithelium, cloning of an enzymatically active and Ca2+/calmodulin-regulated fragment of isoform C and expression in Escherichia coli BL21(DE3)pRIL
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Cloning, Expression, and Purification of RnIP3K-C isoform and of two different recombinant fragments IP3K-C, one comprising the catalytic domain and the calmodulin binding domain, the other comprising only the catalytic domain, are expressed in Escherichi coli BL21(DE3) cells.
entire gene encoding IP3K is cloned and 130fold overexpressed in Escherichia coli BL21(DE3), when induced by isopropyl-beta-D-thiogalactoside
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expression of HA-tagged enzyme in human HEK293 and in COS-7 cells
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expression of mutant cDNA fragment encoding for residues 185-459 of the catalytic domain of isozyme A as unlabeled or selenomethionine-labeled GST fusion proteins in Escherichia coli
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full-length and truncated isoenzyme A is cloned and expressed in HeLa cells, COS-7 cells and primary neuronal cultures
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fusion protein of HsIP3K-C with an N-terminal EGFP tag (EGFP/HsIP3KC) is transiently expressed in NRK52E cells
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gene Itpkb
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gene Itpkc
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IP3K is overexpressed in Escherichia coli
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isoform B is cloned and expressed in pTet-Off-transfected HeLa cells
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stable expression of the C-terminal catalytic domain 3KA-cat30, comprising residues Ser185-Arg459, as N-terminally His-tagged and S-tagged protein in Escherichia coli strain BL21(DE3)
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transient expression of isozyme B in NRK 52E and PC12 cells, DNA and amino acid sequence determination and analysis, expression of isoform B comprising residues 555-934 as Strep-Tactin-tagged enzyme in Escherichia coli strain XL1-Blue, expression of isoform B comprising residues 108-170 as GST-fusion protein in Escherichia coli strain BL21(DE3)
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
overexpression of IP3K-A in DIV 22 hippocampal neurons by infection with adenovirus containing full-length inositol 1,4,5-trisphosphate 3-kinase A leads to a markedly increased number of dendritic structure 18 h after infection, overexpressed GFP-IP3K-A signals are highly localized in the heads of dendritic protrusions
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
K262A
the green fluorescent protein-inositol 1,4,5-trisphosphate 3-kinase A point mutant is generated by site-directed mutagenesis, point mutant lacks kinase activity
D423N
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crystal structure determination and analysis
L217M
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crystal structure determination and analysis
additional information