Information on EC 3.6.3.1 - phospholipid-translocating ATPase

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

EC NUMBER
COMMENTARY hide
3.6.3.1
-
RECOMMENDED NAME
GeneOntology No.
phospholipid-translocating ATPase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + H2O + phospholipid [side 1] = ADP + phosphate + phospholipid [side 2]
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of phosphoric ester
-
-
-
-
transmembrane transport
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
ATP phosphohydrolase (phospholipid-flipping)
A P-type ATPase that undergoes covalent phosphorylation during the transport cycle. The enzyme moves phospholipids such as phosphatidylcholine, phosphatidylserine, and phosphatidylethanolamine from one membrane face to the other ('flippase').
CAS REGISTRY NUMBER
COMMENTARY hide
9000-83-3
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
gerbil
-
-
Manually annotated by BRENDA team
multidrug resistance chinese hamster
-
-
Manually annotated by BRENDA team
strain CCY2811, gene swa4
-
-
Manually annotated by BRENDA team
strain EHY227
-
-
Manually annotated by BRENDA team
strain LMY65, LMY67, LMY69, and LMY94
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-
Manually annotated by BRENDA team
strain sec61
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-
Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
physiological function
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
acetyl phosphate + H2O
acetate + phosphate
show the reaction diagram
-
translocation of phosphatidylserine from the outer to the inner leaflet of resealed erythrocyte ghosts
-
-
?
ATP + H2O
ADP + phosphate
show the reaction diagram
ATP + H2O + 1-palmitoyl-2-(6-(7-nitro-2-1,3-benzoxadiazole)-aminocaproyl)-phosphatidylcholine/out
ADP + phosphate + 1-palmitoyl-2-(6-(7-nitro-2-1,3-benzoxadiazole)-aminocaproyl)-phosphatidylcholine/in
show the reaction diagram
-
internalization through the plasma membrane by the Dnf1p-Lem3p complex
-
-
?
ATP + H2O + 1-palmitoyl-2-(6-(7-nitro-2-1,3-benzoxadiazole)-aminocaproyl)-phosphatidylethanolamine/out
ADP + phosphate + 1-palmitoyl-2-(6-(7-nitro-2-1,3-benzoxadiazole)-aminocaproyl)-phosphatidylethanolamine/in
show the reaction diagram
-
internalization through the plasma membrane by the Dnf1p-Lem3p complex
-
-
?
ATP + H2O + 1-palmitoyl-2-[6-[(7-nitro-2-1,3-benzoxadiazol-4-yl)amino]hexanoyl]-sn-glycero-3-phospho-L-serine/in
ADP + phosphate + 1-palmitoyl-2-[6-[(7-nitro-2-1,3-benzoxadiazol-4-yl)amino]hexanoyl]-sn-glycero-3-phospho-L-serine/out
show the reaction diagram
-
-
-
?
ATP + H2O + 7-nitrobenz-2-oxa-1,3-diazol-4-yl-phosphatidylserine/in
ADP + phosphate + 7-nitrobenz-2-oxa-1,3-diazol-4-yl-phosphatidylserine/out
show the reaction diagram
ATP + H2O + asolectin/out
ADP + phosphate + asolectin/in
show the reaction diagram
-
-
-
-
?
ATP + H2O + carboxyfluoresceinyl-phosphatidylserine/in
ADP + phosphate + carboxyfluoresceinyl-phosphatidylserine/out
show the reaction diagram
ATP + H2O + cholera toxin/in
ADP + phosphate + cholera toxin/out
show the reaction diagram
-
through ATP hydrolysis and an interaction with the cytoplasmic membrane protein EpsL, EpsE supports secretion of cholera toxin across the outer membrane
-
-
?
ATP + H2O + coumaryl-phosphatidylserine/in
ADP + phosphate + coumaryl-phosphatidylserine/out
show the reaction diagram
ATP + H2O + galactocerebroside/out
ADP + phosphate + galactocerebroside/in
show the reaction diagram
-
-
-
-
?
ATP + H2O + H33342/out
ADP + phosphate + H33342/in
show the reaction diagram
-
-
-
-
?
ATP + H2O + lactocerebroside/out
ADP + phosphate + lactocerebroside/in
show the reaction diagram
-
-
-
-
?
ATP + H2O + Lipid A/in
ADP + phosphate + Lipid A/out
show the reaction diagram
-
-
-
?
ATP + H2O + lysophosphatidylcholine/out
ADP + phosphate + lysophosphatidylcholine/in
show the reaction diagram
-
lysophosphatidylcholine is a bona fide biological substrate transported by the yeast plasma membrane ATPases, Dnf1p and Dnf2p, in consort with a second protein Lem3p
-
-
?
ATP + H2O + lysophosphatidylethanolamine/out
ADP + phosphate + lysophosphatidylethanolamine/in
show the reaction diagram
-
lysophosphatidylethanolamine is a bona fide biological substrate transported by the yeast plasma membrane ATPases, Dnf1p and Dnf2p, in consort with a second protein Lem3p
-
-
?
ATP + H2O + miltefosine/in
ADP + phosphate + miltefosine/out
show the reaction diagram
ATP + H2O + N-methyl-dilauroylphosphatidyl-DL-serine/out
ADP + phosphate + N-methyl-dilauroylphosphatidyl-DL-serine/in
show the reaction diagram
-
rapid change of cell morphology from echinocyte to stomatocyte
-
-
?
ATP + H2O + phosphatidylcholine/in
ADP + phosphate + phosphatidylcholine/out
show the reaction diagram
ATP + H2O + phosphatidylcholine/in
ADP + phosphate + phosphatidylethanolamine/out
show the reaction diagram
ATP + H2O + phosphatidylcholine/out
ADP + phosphate + phosphatidylcholine/in
show the reaction diagram
-
-
-
-
?
ATP + H2O + phosphatidylethanolamine/in
ADP + phosphate + phosphatidylethanolamine/out
show the reaction diagram
ATP + H2O + phosphatidylethanolamine/out
ADP + phosphate + phosphatidylethanolamine/in
show the reaction diagram
ATP + H2O + phosphatidylserine/in
ADP + phosphate + phosphatidylserine/out
show the reaction diagram
ATP + H2O + phosphatidylserine/out
ADP + phosphate + phosphatidylserine/in
show the reaction diagram
ATP + H2O + phosphoethanolamine/in
ADP + phosphate + phosphoethanolamine/out
show the reaction diagram
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low activity
-
-
?
ATP + H2O + phospholipid/in
ADP + phosphate + phospholipid/out
show the reaction diagram
ATP + H2O + rhodamine 123/out
ADP + phosphate + rhodamine 123/in
show the reaction diagram
-
-
-
-
?
ATP + H2O + sn-1,2-dilauroylphosphatidyl-D-serine/out
ADP + phosphate + sn-1,2-dilauroylphosphatidyl-D-serine/in
show the reaction diagram
-
rapid change of cell morphology from echinocyte to stomatocyte
-
-
?
ATP + H2O + sn-1,2-dilauroylphosphatidyl-L-serine/out
ADP + phosphate + sn-1,2-dilauroylphosphatidyl-L-serine/in
show the reaction diagram
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rapid change of cell morphology from echinocyte to stomatocyte
-
-
?
ATP + H2O + sn-2,3-dilauroylphosphatidyl-L-serine/out
ADP + phosphate + sn-2,3-dilauroylphosphatidyl-L-serine/in
show the reaction diagram
-
low activity, slow change of cell morphology from echinocyte to stomatocyte
-
-
?
ATP + H2O + sphingomyelin/out
ADP + phosphate + sphingomyelin/in
show the reaction diagram
-
-
-
-
?
ATP + H2O + tetramethylrosamine/out
ADP + phosphate + tetramethylrosamine/in
show the reaction diagram
-
-
-
-
?
ATP + H2O + verapamil/out
ADP + phosphate + verapamil/in
show the reaction diagram
-
highest activity
-
-
?
dATP + H2O
dADP + H2O
show the reaction diagram
GTP + H2O
GDP + phosphate
show the reaction diagram
-
at 2 mM, 40% of the activity with 2 mM ATP
-
-
?
p-nitrophenyl phosphate + H2O
p-nitrophenol + phosphate
show the reaction diagram
-
translocation of phosphatidylserine from the outer to the inner leaflet of resealed erythrocyte ghosts
-
-
?
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 + H2O
ADP + phosphate
show the reaction diagram
ATP + H2O + miltefosine/in
ADP + phosphate + miltefosine/out
show the reaction diagram
-
LdMT and LdRos3 form part of the same translocation machinery that determines flippase activity and miltefosine sensitivity in Leishmania
-
-
?
ATP + H2O + phosphatidylethanolamine/in
ADP + phosphate + phosphatidylethanolamine/out
show the reaction diagram
ATP + H2O + phosphatidylethanolamine/out
ADP + phosphate + phosphatidylethanolamine/in
show the reaction diagram
ATP + H2O + phosphatidylserine/in
ADP + phosphate + phosphatidylserine/out
show the reaction diagram
ATP + H2O + phosphatidylserine/out
ADP + phosphate + phosphatidylserine/in
show the reaction diagram
ATP + H2O + phosphoethanolamine/in
ADP + phosphate + phosphoethanolamine/out
show the reaction diagram
-
low activity
-
-
?
ATP + H2O + phospholipid/in
ADP + phosphate + phospholipid/out
show the reaction diagram
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1-monoolein
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0.5 mM significantly reduce the P-glycoprotein protein expression, 0.1 mM of 1-monoolein does not have any significant effect on the expression of P-glycoprotein
1-monostearin
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0.5 mM significantly reduce the P-glycoprotein protein expression, 0.1 mM of 1-monostearin does not have any significant effect on the expression of P-glycoprotein
4-hydroxy-2-nonenal
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-
AlFx
trapping of MsbA with 0.8 mM AlFx results in incomplete inhibition of activity
azide
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-
BeFx
trapping of MsbA with 0.8 mM BeFx results in incomplete inhibition of activity
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Cd2+
-
-
cholesterol
-
the transport activity of P-glycoprotein decreases by about 20% cholesterol
cyclosporin A
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inhibition of phospholipid flipping
Diamide
-
-
DTNB
-
-
elaiophylin
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partial inhibition of Mg+-ATPase activity, translocation of phosphatidylserine is almost completely abolished
eosin Y
-
almost complete inhibition of both Mg2+-ATPase activity and translocation
Kes1p
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Kes1p represses the flippase activity of Drs2p in trans-Golgi network membranes
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lipid A
lipid A inhibits translocation by approx. 30% at 0.01-0.04 mg/ml
methyl-beta-cyclodextrin
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transport of 0.001 mM tetramethylrosamine is essentially abolished by 1 mM methyl-beta-cyclodextrin, and transport of 0.005 mM H33342 is almost completely inhibited by 5 mM methyl-beta-cyclodextrin
orthovanadate
-
-
phosphate analogue Vi
the presence of 0.1-0.2 m M Vi inhibits flippase activity of MsbA by approx. 50%
pyridyldithioethylamine
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-
sulfhydryl reagents
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-
suramin
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competitive inhibitor of ATP towards both Mg2+-ATPase activity and aminophospholipid translocation. Inhibition of translocation occurs at higher inhibitor concentration than the inhibition of Mg2+-ATPase activity
vanadate
vanadyl ion
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inhibits the enzyme from the extracellular surface
verapamil
vinblastine
-
inhibition of phospholipid flipping
additional information
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1-aminonaphthalene-8-sulfonate
-
stimulates
1-heptanesulfonate
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stimulates
2,4-Dinitrophenol
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stimulates, stimulation is prevented by 0.2-1.0 mM Ca2+
4-Aminosalicylate
-
stimulates
acidic phospholipids
-
acidic phospholipids, specifically cardiolipin, bind the co-purified EpsE/cyto-EpsL complex and stimulate its ATPase activity 30-130fold, whereas the activity of EpsE alone is unaffected. Removal of the last 11 residues, residues 243-253, from cyto-EpsL prevented cardiolipin binding as well as stimulation of the ATPase activity of EpsE
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ALIS protein
-
-
-
Benzoate
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stimulates
Brij
-
stimulates
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cardiolipin
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binds the co-purified EpsE/cyto-EpsL complex and stimulates its ATPase activity 30-130fold, whereas the activity of EpsE alone is unaffected. Removal of the last 11 residues, residues 243-253, from cyto-EpsL prevents cardiolipin binding as well as stimulation of the ATPase activity of EpsE
CDC50 proteins
CDC50A and CDC50B, cloning and expression of the abut 60 kDa proteins, required for full translocase activity, 2.5-5fold activition, proteins are pivotal factors in the trafficking of ATP8B1 to the plasma membrane and thus may be essential determinants of ATP8B1-related disease, physical interacion with ATP8B1, overview
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CDC50A
-
-
-
cholate
cholesterol
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the transport activity of P-glycoprotein is elevated by about 40% cholesterol
dithiothreitol
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partially stimulates
glycerophosphoserine
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prevents transport of phosphatidylserine
glycocholate
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slightly increases the efflux of phospholipids and cholesterol from cells
Kdo2-lipid A
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-
p-hydroxyphenylacetate
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stimulates
p-nitrophenol
-
stimulates
phenylacylbromide
-
-
Phenylglyoxal
-
-
phosphatidylethanolamine
phosphatidylglycerol
-
slight activation
phosphatidylhomoserine
-
slight activation
phosphatidylhydroxypropionate
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slight activation
phosphatidylserine
Phospholipids
-
The activity of purified MsbA is dependent upon the presence of phospholipids.
protein Drs2p
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coupled to the enzyme, required for the translocation of phospholipids from the luminal leaflet of the membrane to the cytosolic leaflet in the trans-Golgi network, overview
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salicylate
-
stimulates
SDS
-
stimulates
sulfanilamide
-
stimulates
Sulfanilic acid
-
stimulates
taurocholate
-
increases the efflux of phospholipids and cholesterol from cells, the taurocholate monomer plays an important role in ABCB4-mediated lipid secretion
Tetradecyltrimethylammonium bromide
-
stimulates
thrombin
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induces platelet phosphatidylserine exposure, inhibited by c7E3 or SR121566
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Triton X-100
-
stimulates
verapamil
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a P-glycoprotein modulator known to stimulate ATPase activity in a biphasic manner at micromolar concentrations
additional information
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.03 - 1.31
acetyl phosphate
0.016 - 2.215
ATP
1.17 - 1.46
p-nitrophenyl phosphate
additional information
additional information
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.002 - 0.004
-
activity of purified, detergent-solubilized MsbA
0.054
purified native enzyme, pH 7.5, 37C
2.3
-
Atp8a1 in Sf21 cell lysate, in absence of phosphatidylserine
4.9
-
Atp8a1 in Sf21 cell lysate, in presence of phosphatidylserine
35
purified native enzyme reconstituted in lipid vesicles, pH 7.5, 37C
45 - 55
purified lipid-stimulated enzyme, pH 7.5, 37C
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7 - 9
with substrate phosphatidylserine
7 - 8.5
-
-
7
-
assay at
8
with substrate phosphatidylethanolamine
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
-
below 4C transport of phospholipids is inhibited in both erythrocytes and fibroblasts, warming to 7C activates transport in fibroblasts
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
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epithelial cells
Manually annotated by BRENDA team
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adult ventricular
Manually annotated by BRENDA team
-
neonatal rat cardiomyoblasts
Manually annotated by BRENDA team
high distribution in pancreas
Manually annotated by BRENDA team
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expression analysis of isozymes Atp8a1 and flippase in reticulocytes, overview
Manually annotated by BRENDA team
high distribution in small intestine
Manually annotated by BRENDA team
ATP8B3 is expressed only in germ cells, especially in haploid spermatids
Manually annotated by BRENDA team
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
localized in developing acrosomes
-
Manually annotated by BRENDA team
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AP-1 is required, and GGA proteins, Golgi localized, gamma-ear containing, Arf-binding proteins acting as chlatrin adaptos, are dispensable, for efficient exclusion of Drs2p from exocytic vesicles targeted to the plasma membrane
Manually annotated by BRENDA team
-
EpsE is a cytoplasmic component of the type II secretion system
Manually annotated by BRENDA team
ATP8B1 in absence of CDC50A and CDC50
Manually annotated by BRENDA team
-
Cdc50p and Drs2p are localized to the trans-Golgi network and late endosome
Manually annotated by BRENDA team
additional information
PDB
SCOP
CATH
ORGANISM
UNIPROT
Mycobacterium tuberculosis (strain CDC 1551 / Oshkosh)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
160000
SDS-PAGE
170000
-
SDS-PAGE
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
heterodimer
homodimer
2 * 65000, SDS-PAGE
additional information
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycoprotein
-
analysis of glycosylation of ABCB4, cleavage by endoglycosidase H or peptide N-glycosidase F, overview
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
the enzyme is very instable and rapidly degenerated during purification
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
4C, 35 days, stored erythrocytes show pH-dependently reduced flippase activity and ATP depletion, upon ATP supply the activity can be restored, overview
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ATP8A1 from chromaffin granules
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concanavalin A-Sepharose 4B affinity column chromatography and Sephadex G50 gel filtration
-
native Atp8a2 1887fold from photoreceptor outer segments by immunoaffinity chromatography
native enzyme from chromaffin granules
-
native enzymes partially by subcellular fractionation
-
Ni-NTA column chromatography
partial
-
partially by microsome preparation
-
partially from erythrocytes
-
recombinant enzyme from HEK-293 cells by immunoaffinity chromatography
recombinant His-tagged Atp8a1 from Spodoptera frugiperda Sf21 cells by microsome preparation and nickel affinity chromatography
-
recombinant wild-type and mutant enzymes
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ATP10C
-
Creation and study of strains lacking Dnf1, Dnf2 or both Dnf1 and Dnf2. The double mutant is defective in the inward translocalisation of NBD-labeled phosphatidylethanolamine, phosphatidylserine and phosphatidylcholine across the plasmamembrane. The loss of both Dnf1 and Dnf2 leads to an increased cell surface exposure of endogenous aminophospholipids. The double mutant is sensitive to low concentrations of Co2+, Ni2+, Zn2+, and Mn2+
-
expressed in COS-7 cells
expressed in Saccharomyces cerevisiae strain w303-1a
expressed in UPS-1 cells
expression of GFP-tagged Drs2p and HA-tagged Drf1p wild-type and mutant enzymes in Saccharomyces cerevisiae, subcloning in Escherichia coli strains XL1-Blue and DH5alpha
-
expression of His-tagged Atp8a1 in Spodoptera frugiperda Sf21 cells using the baculovirus transfection system
-
gene atp8a2, DNA and amino acid sequence determination and analysis, expression in HEK-293 cells
gene atp8a2, DNA and amino acid sequence determination and analysis, expression of 1D4-tagged Atp8a2 in HEK-293 cells
gene swa4, DNA and amino acid sequence determination and anaylsis, comparison to the allelic gene cdc50
-
genes DRS2, DNF1 and DNF2, and DNF3, expression of wild-type and mutant enzymes, overview
-
isolated stable expression of N-terminally GFP-tagged ATP8B1 in UPS-1 cells, a nonpolarized CHO-K1 mutant cell line with a defect in the nonendocytic uptake of the NBD-PS analogue, by lentiviral transduction leading to protein localization in the endoplasmic reticulum, co-expression with N-terminally HA-tagged CDC50 proteins results in relocalization of ATP8B1 from the endoplasmic reticulum to the plasma membrane, natural phosphatidylserine exposure in the outer leaflet of the plasma membrane was reduced by 17%-25% in cells coexpressing ATP8B1 and CDC50 proteins in comparison with cells expressing ATP8B1 alone, coexpression of ATP8B1 and CDC50A in WIF-B9 cells resulted in colocalization of both proteins in the canalicular membrane, overview
LdMT is cloned by rescuing the resistance phenotype of the M-40 R cell line overexpression in Leishmania tarentolae
-
stable expression of ABCB4 in HEK-293 cells
-
the gene MsbA is cloned into pET28b behind the T7 promoter in-frame with an N-terminal His6tag
-
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
coexpression of CDC50A increased the total cellular expression level of flippase ATP8A1
-
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
E371Q
affinity to phosphatidylserine similar to wild-type
F354A
affinity to phosphatidylserine similar to wild-type
F88A
tendency for inhibition at high substrate concentration
I115A
biphasic phosphatidylserine concentration dependence with inhibition at the highest concentration
I362A
increase in the apparent affinity for phosphatidylserine
I364A
80% of wild-type activity
I364E
3-5fold reduction in affinity for phosphatidylserine
I364F
3-5fold reduction in affinity for phosphatidylserine
I364M
3-5fold reduction in affinity for phosphatidylserine
I364Q
activation phase is followed by an inhibition phase at high phosphatidylserine concentration
I364S
30% of wild-type activity
K374A
affinity to phosphatidylserine similar to wild-type
L112A
mutation does not appreciably affect Vmax, the apparent affinities for the substrates, or the phosphorylation rate
L361A
affinity to phosphatidylserine similar to wild-type
L367A
significant reduction in affinity to phosphatidylserine
N359A
dramatic reduction of Vmax to 9-11% of wild-type
N360A
increase in the apparent affinity for phosphatidylserine
P363A
complete loss of activity
S365A
significant reduction in affinity to phosphatidylserine
T369A
increase in the apparent affinity for phosphatidylserine
V906A
pronounced inhibition at high phosphatidylethanol concentration with only a slight inhibition at high phosphatidylserine concentration
Y358A
affinity to phosphatidylserine similar to wild-type
A270T
-
mutant and wild-type enzymes have similar activities at 30C, but the mutant activity is decreased significant at 42C
I376M
-
missense mutation identified in a patient with cerebellar ataxia, mental retardation and dysequilibrium syndrome. The mutation lies in a highly conserved C-terminal transmembrane region of E1 E2 ATPase domain
K1075M
-
site-directed mutagenesis in the Walker A motif results in an inactive mutant
K435M
-
site-directed mutagenesis in the Walker A motif results in an inactive mutant
G308V
Atp8b1 protein is virtually undetectable in G308V mouse liver, the G308V substitution results in an instable protein that cannot exit the endoplasmic reticulum and is broken down by the proteasome
F511L
-
the mutant has a specific activity that is 35% that of wild type Drs2
F511Y
-
the mutant retains wild type activity, the substitution in Drs2 specifically abrogates phosphatidylserine recognition by this flippase causing phosphatidylserine exposure on the outer leaflet of the plasma membrane without disrupting phospatidylethanolamine asymmetry
I1235F
-
mutation increases the overall activity of Dnf1 for all substrates and causes partial loss of specificity for glycerophospholipid
Y618F
-
acquisition of the phosphatidylserine substrate maps to a Tyr618Phe substitution in transmembrane segment 4 of Dnf1. The rate of 7-nitrobenz-2oxa-1,3-diazol-4-yl phospholipid uptake by Dnf1 Y618F is comparable to wild type Dnf1
additional information
Renatured/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
preparation of proteoliposomes for enzyme reconstitution using microsome and liposomes, overview
-
purified Atp8a2 is reconstituted into liposomes containing fluorescent-labeled phosphatidylserine to measure the ability of Atp8a2 to flip phosphatidylserine across the lipid bilayer. 30% of the reconstituted Atp8a2 is inaccessible to trypsin and correspondingly ATP
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
assay for phospholipid flippase activities of plasma membrane-localized P4-ATPases using human cell lines stably expressing isoforms ATP8B1, ATP8B2, ATP11A, and ATP11C; assay for phospholipid flippase activities of plasma membrane-localized P4-ATPases using human cell lines stably expressing isoforms ATP8B1, ATP8B2, ATP11A, and ATP11C; assay for phospholipid flippase activities of plasma membrane-localized P4-ATPases using human cell lines stably expressing isoforms ATP8B1, ATP8B2, ATP11A, and ATP11C; assay for phospholipid flippase activities of plasma membrane-localized P4-ATPases using human cell lines stably expressing isoforms ATP8B1, ATP8B2, ATP11A, and ATP11C
medicine
additional information
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Drs2p deficiency causes a markedly increased rate of cholesterol transport from the plasma membrane to the endoplasmic reticulum and redistribution of endogenous ergosterol to intracellular membranes
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