Information on EC 3.1.3.64 - phosphatidylinositol-3-phosphatase

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

EC NUMBER
COMMENTARY hide
3.1.3.64
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RECOMMENDED NAME
GeneOntology No.
phosphatidylinositol-3-phosphatase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
1-phosphatidyl-1D-myo-inositol 3-phosphate + H2O = 1-phosphatidyl-1D-myo-inositol + phosphate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of phosphoric ester
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
3-phosphoinositide degradation
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phosphatidylinositol biosynthesis I (bacteria)
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Inositol phosphate metabolism
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Metabolic pathways
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SYSTEMATIC NAME
IUBMB Comments
1-phosphatidyl-1D-myo-inositol-3-phosphate 3-phosphohydrolase
This enzyme still works when the 2,3-bis(acyloxy)propyl group is removed, i.e., it hydrolyses Ins(1,3)P2 to Ins-1-P.
CAS REGISTRY NUMBER
COMMENTARY hide
122653-77-4
formerly, EC 3.1.3.65
124248-47-1
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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D3HMN4
UniProt
Manually annotated by BRENDA team
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D3HMN4
UniProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
metabolism
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
1-hexanoyl-2-lysophosphatidyl-1D-myo-inositol 3,5-bisphosphate + H2O
1-hexanoyl-2-lysophosphatidyl-1D-myo-inositol 5-phosphate + phosphate
show the reaction diagram
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-
-
?
1-phosphatidyl-1D-myo-inositol 3,5-bisphosphate + H2O
1-phosphatidyl-1D-myo-inositol 5-phosphate + phosphate
show the reaction diagram
1-phosphatidyl-1D-myo-inositol 3-phosphate + H2O
1-phosphatidyl-1D-myo-inositol + phosphate
show the reaction diagram
hexanoylglyceryl 1-phosphatidyl-1D-myo-inositol 3-phosphate + H2O
hexanoylglyceryl 1-phosphatidyl-1D-myo-inositol + phosphate
show the reaction diagram
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?
inositol 1,3-bisphosphate + H2O
inositol 1-phosphate + phosphate
show the reaction diagram
phosphatidylinositol 3,4-bisphosphate + H2O
phosphatidylinositol 4-phosphate + phosphate
show the reaction diagram
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?
phosphatidylinositol 3,5-bisphosphate + H2O
phosphatidylinositol 5-phosphate + phosphate
show the reaction diagram
phosphatidylinositol 3-phosphate + H2O
phosphatidylinositol + phosphate
show the reaction diagram
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
1-phosphatidyl-1D-myo-inositol 3,5-bisphosphate + H2O
1-phosphatidyl-1D-myo-inositol 5-phosphate + phosphate
show the reaction diagram
1-phosphatidyl-1D-myo-inositol 3-phosphate + H2O
1-phosphatidyl-1D-myo-inositol + phosphate
show the reaction diagram
inositol 1,3-bisphosphate + H2O
inositol 1-phosphate + phosphate
show the reaction diagram
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type I enzyme may regulate inositol 1,3-bisphosphate levels in cells and is designated inositol-polyphosphate 3-phosphatase
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?
phosphatidylinositol 3,5-bisphosphate + H2O
phosphatidylinositol 5-phosphate + phosphate
show the reaction diagram
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physiologically relevant substrate
product could activate enzyme by a positive feedback mechanism
?
phosphatidylinositol 3-phosphate + H2O
phosphatidylinositol + phosphate
show the reaction diagram
additional information
?
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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no metal ion requirement
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Ca2+
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in the millimolar range, 1 mM: 50-75% inhibition
Mg2+
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in the millimolar range, slightly less potent than Ca2+
type III PI3K regulatory subunit hVps15
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i.e. p150, MTMR2 binding results in inhibition of phosphatase activity and in diminution of hVps34-Rab7 interaction important for PI3P synthesis
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VO43-
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0.1 mM: 15% inhibition, 0.5 mM: 95% inhibition
Zn2+
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0.5 mM: 90% inhibition
additional information
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
EGTA
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activation, optimum at 1.5-3 mM
myotubularin-related protein 9
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myotubularin-related protein 9 increases the binding of myotubularin-related protein 6 to phospholipids without changing the lipid binding profile, myotubularin-related protein 9 increases the 3-phosphatase activity of myotubularin-related protein 6 up to 6fold, myotubularin-related protein 9 potentiates the effects of myotubularin-related protein 6 on apoptosis
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octyl glucoside
phosphatidylinositol 3-phosphate
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substrate is a weak allosteric activator of MTM1 capable of binding at a regulatory site on the monomeric enzyme, no physiological activator
phosphatidylinositol 4-phosphate
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phosphatidylinositol 4-phosphate increases myotubularin-related protein 6 activity in the presence of myotubularin-related protein 9 in 100% phosphatidylserine
phosphatidylinositol 5-phosphate
phosphatidylserine
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myotubularin-related protein 6 activity increases 1.8fold in both 50% and 100% phosphatidylserine
additional information
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myotubularin-related protein 6 is activated up to 28fold in the presence of phosphatidylserine liposomes, myotubularin-related protein 6 activity in the presence of myotubularin-related protein 9 and assayed in phosphatidylserine liposomes increases 84fold
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0008 - 0.0037
Inositol 1,3-bisphosphate
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.8
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assay at
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
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satellite cells of dorsal root ganglia
Manually annotated by BRENDA team
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fibroblast cells
Manually annotated by BRENDA team
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detected in vivo in axons in peripheral nerves
Manually annotated by BRENDA team
adapter subunit 3-PAP mRNA, highest abundance in liver, kidney, lung and placenta, 2 transcripts of 5.5 and 2.5 kb
Manually annotated by BRENDA team
isolation of adapter subunit 3-PAP immunoprecipitates
Manually annotated by BRENDA team
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ventral roots of spinal cord
Manually annotated by BRENDA team
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contains a spliced isoform
Manually annotated by BRENDA team
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
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condensed in the perinuclear region in a microtubule-dependent manner. Rab1B regulates the localization of MTMR6
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Manually annotated by BRENDA team
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MTMR2 is localized to excitatory synapses of central neurons, localization in synaptic membrane, cytosol, and vesicles
Manually annotated by BRENDA team
additional information
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
65000
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1 * 65000 + 1 * 78000, type II 3-phosphatase, SDS-PAGE; 2 * 65000, type I 3-phosphatase, SDS-PAGE
71000
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SDS-PAGE
78000
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1 * 65000 + 1 * 78000, type II 3-phosphatase, SDS-PAGE
86000
x * 86000 + x * ?, adapter subunit 3-PAP of phosphatidylinositol 3-phosphate phosphatase, forms a complex with a catalytic subunit of unknown MW, predicted from the amino acid sequence
110000
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type I 3-phosphatase, gel filtration
115000
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SDS-PAGE
147000
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type II 3-phosphatase, gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
heterodimer
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1 * 65000 + 1 * 78000, type II 3-phosphatase, SDS-PAGE; the 65 kDa subunit of the type I and II enzyme appear to be identical, based on co-migration in SDS-polyacrylamide gels and peptide mapping, the 78 kDa subunit of the type II enzyme may be a regulatory subunit
homodimer
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2 * 65000, type I 3-phosphatase, SDS-PAGE; the 65 kDa subunit of the type I and II enzyme appear to be identical, based on co-migration in SDS-polyacrylamide gels and peptide mapping
monomer
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MTM1 and MTMR6, catalytically inactive in absence of substrate, binding of substrate and of the allosteric activator phosphatidylinositol 5-phosphate triggers oligomerization and thus activates enzyme
oligomer
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MTM1 and MTMR6, catalytically inactive monomer in absence of substrate, binding of substrate and of the allosteric activator phosphatidylinositol 5-phosphate triggers oligomerization and thus activates enzyme
additional information
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phosphoprotein
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ERK1/2 is the kinase responsible for phosphorylating MTMR2 at position Ser58, which suggests that the endosomal targeting of MTMR2 is regulated through an ERK1/2 negative feedback mechanism
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
crystal structure of SidP orthologue from Legionella longbeachae reveals that this unique PI-3-phosphatase is composed of three distinct domains: a large catalytic domain, an appendage domain that is inserted into the N-terminal portion of the catalytic domain, and a C-terminalalpha-helical domain
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-90°C, frozen in liquid N2, at least 1 month, stable
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
anti-FLAG affinity column chromatography
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partial, two enzyme forms: type I and type II 3-phosphatase
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recombinant FLAG-tagged and His-tagged wild-type and mutant enzymes from Escherichia coli lysates
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recombinant GST-tagged enzyme from Escherichia coli strain DH5alpha by glutathione affinity chromatography
recombinant MTM1 and MTMR6
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cDNA encoding the adapter subunit 3-PAP of phosphatidylinositol 3-phosphate 3-phosphatase is cloned, sequenced and characterized, expression of 3-PAP in SF9 insect cells
cDNAs encoding MTM1 and MTMR6 are cloned and expressed in SF9 insect cells, MTM1 is also expressed in yeast
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DNA and amino acid sequence analysis, phylogenetic analysis, coexpression of FLAG-tagged MTMR2 and MTMR13 in HEK-293A cells
expressed as an N-terminal FLAG-tagged fusion construct
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expressed in Escherichia coli as a His-tagged fusion protein
expressed in Sf9 insect cells, HEK-293 TRex cells, and in HeLa cells
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expression of GST-tagged enzyme in Escherichia coli strain DH5alpha, overexpression in COS-7 cells, HEK-293 cells, L6 cells, C2C12 cells, and Jurkat with 10-15% expression level in all cases
expression of wild-type myotubularin-related phosphatase in A-549 cells, overexpression of a dominant-negative inactive mutant of myotubularin-related phosphatase 3, that partially localizes to autophagosomes, and PtdIns3P and two autophagy-related PtdIns3P-binding proteins, GFPDFCP1 and GFP-WIPI-1alpha, in A-549 cells, the recombinant enzyme accumulates at sites of autophagosome formation
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overexpression in COS-7 cells
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overexpression of wild-type MTMR2 in HeLa cells predominantly located to the cytosolic/perinuclear region, recombinant expression of FLAG-tagged and of His-tagged wild-type and mutant enzymes in Escherichia coli lysates
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stable coexpression of human FLAG-tagged Ca2+-activated K+ channel KCa3.1 with GFP-tagged or FLAG-tagged MTMR6 in CHO cells
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C330S
strong decrease in enzymatic activity (27% of wild type activity), mutant displays a similar localization pattern to the wild-type construct
C336S
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inactive MTMR6 active site mutant
C413S
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a dominant-negative inactive mutant of myotubularin-related phosphatase 3. The distribution of MTMR3C413S substantially overlaps with both autophagy-related PtdIns3P-binding proteins GST-2xFYVE and GFP-LC3, demonstrating that the amount of PtdIns3P is focally increased in areas of MTMR3C413S accumulation on autophagosomes
C417S
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site-directed mutagenesis, a catalytically inactive MTMR2 variant
D278A
site-directed mutagenesis, inactive, substrate-trapping mutant
G103E
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mutations in the PH-GRAM domain affect both binding to the membrane and the phosphatase activity, as demonstrated for the G103E mutation
R114A
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MTM1 mutant, conserved residue, decreased response to the allosteric activator phosphatidylinositol 5-phosphate
R184G
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MTM1 mutant with reduced activity
R220A
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inactive MTM1 mutant, conserved residue
R241L
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inactive MTM1 mutant
R421Q
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MTM1 mutant with reduced activity
R69C
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a disease-causing mutation of MTM1 falling within a putative pleckstrin homology domain, decreased response to the allosteric activator phosphatidylinositol 5-phosphate due to a reduced affinity for the activator, mutant with reduced activity due to a mutation in the allosteric regulatory site
S58A/C417S
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site-directed mutagenesis, a catalytically inactive MTMR2 variant, which shows strong co-localization with markers for early endosomes including the PI(3)P marker generated from EEA1 and Rab5
S58A/C417S/S631A
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catalytically inactive variant. Loss of catalytic activity (S58A/C417S/S631A) of dephosphorylated MTMR2 results in partial colocalization with PI(3)P endosomes. In contrast to double mutant S58A/S631A triple mutant S58A/C417S/S631A containing an additional mutated catalytic residue do not show enlarged viscles being positive for both MTMR2 and APPL1. This underlines that that the phosphatase activity of MTMR2 contributes to the observed enlargement of MTMR2/APPL1 positive vesicles
S58A/S631A
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dephosphorylation at Ser58 and Ser631 in double mutant S58A/S631A prevents localization to PI(3)P-rich endosomes. MTMR2 localization shift upon administration of MAPK inhibitors is recapitulated in double mutant S58A/S631A. Expression of this double mutant leads to a more sustained and pronounced increase in ERK1/2 activation compared with mutant S58A. Further analysis of single mutant S58A and double mutant S58A/S631A demonstrate that it is the phosphorylation status of Ser58 that regulates general endosomal binding and that the phosphorylation status of Ser631 mediates the endosomal shuttling between Rab5 and APPL1 subtypes
S58E
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site-directed mutagenesis, a phosphorylation-deficient mutant, which retains full catalytic activity toward PI(3)P and PI(3,5)P2 as compared to wild-type MTMR2. The mutant enzyme displays a similar localization pattern as wild-type in the cytoplasm
Y462C
decreased enzymatic activity (84% of wild type activity), mutant displays a similar localization pattern to the wild-type construct. This mutant is found as a non-conservative heterozygous Y462C missense in a patient suffering from centronuclear myopathy
C554S
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mutant does not show any activity
D559N
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mutant does not show any activity
R560K
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mutant does not show any activity
E276X
another animal model for Charcot-Marie-Tooth type 4B1 is produced by introducing the E276X mutation in exon 9. Phosphatase activity is inactivated toward PtdIns3P and PtdIns(3,5)P2. Nerve morphology in these mice is similar to that observed in Mtmr2-null mice, although a huge variability in the number of myelin outfoldings between different mice is noted. In both mutants, Mtmr2-null and E276X, the dysmyelinating phenotype is less severe than that observed in human Charcot-Marie-Tooth type 4B1
Q426X
phosphatase activity is inactivated toward PtdIns3P and PtdIns(3,5)P2 in vitro
Q482X
phosphatase activity is inactivated toward PtdIns3P and PtdIns(3,5)P2 in vitro
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
medicine