Information on EC 3.1.1.64 - retinoid isomerohydrolase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
3.1.1.64
-
RECOMMENDED NAME
GeneOntology No.
retinoid isomerohydrolase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
an all-trans-retinyl ester + H2O = 11-cis-retinol + a fatty acid
show the reaction diagram
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-
-
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of carboxylic ester
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
the visual cycle I (vertebrates)
-
-
Retinol metabolism
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SYSTEMATIC NAME
IUBMB Comments
all-trans-retinyl ester acylhydrolase, 11-cis retinol forming
This enzyme, which operates in the retinal pigment epithelium (RPE), catalyses the cleavage and isomerization of all-trans-retinyl fatty acid esters to 11-cis-retinol, a key step in the regeneration of the visual chromophore in the vertebrate visual cycle [4]. Interaction of the enzyme with the membrane is critical for its enzymic activity [6].
CAS REGISTRY NUMBER
COMMENTARY hide
106389-24-6
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
no activity in cephalochordata
-
-
-
Manually annotated by BRENDA team
no activity in tunicata
-
-
-
Manually annotated by BRENDA team
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UniProt
Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
malfunction
metabolism
-
the enzyme catalyzes the key reaction in the retinoid visual cycle
physiological function
additional information
-
key residues determining the isomerization product specificity of the enzyme are Tyr58, Phe103, and Leu133
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
11-cis-retinol + a fatty acid
all-trans-retinyl ester + H2O
show the reaction diagram
-
the reverse isomerization occurs in vivo
-
-
r
all-trans-retinol
11-cis-retinol
show the reaction diagram
all-trans-retinyl acetate + H2O
all-trans-retinol + acetate
show the reaction diagram
-
-
-
-
?
all-trans-retinyl ester + H2O
11-cis-retinal + a fatty acid
show the reaction diagram
-
-
-
-
?
all-trans-retinyl ester + H2O
11-cis-retinol + 13-cis-retinol + a fatty acid
show the reaction diagram
intrinsic substrate for isoform RPE65c
-
-
?
all-trans-retinyl ester + H2O
11-cis-retinol + a fatty acid
show the reaction diagram
all-trans-retinyl palmitate + H2O
11-cis-retinol + palmitate
show the reaction diagram
all-trans-retinyl palmitate + H2O
all-trans-retinol + palmitate
show the reaction diagram
an all-trans retinyl ester + H2O
13-cis-retinol + a fatty acid
show the reaction diagram
specific substrate
-
-
?
an all-trans-retinyl ester + H2O
11-cis-retinol + a fatty acid
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
11-cis-retinol + a fatty acid
all-trans-retinyl ester + H2O
show the reaction diagram
-
the reverse isomerization occurs in vivo
-
-
r
all-trans-retinyl ester + H2O
11-cis-retinol + a fatty acid
show the reaction diagram
an all-trans-retinyl ester + H2O
11-cis-retinol + a fatty acid
show the reaction diagram
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1,10-phenanthroline
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dose-dependent inhibition
10-N-acetamidodecyl chloromethyl ketone
2,2'-bipyridine
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dose-dependent inhibition
2,2'-dipyridyl
-
-
2,2-dimethyl-4-phenyl-2H-imidazole-1-oxide
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strong inhibition of isomerohydrolase activity (32% at 0.2 mM) in vitro
2-(2-carboxyethyl)-2-methyl-4-phenyl-2H-imidazole-1-oxide
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moderate inhibitory effect at 0.2 mM
all-trans-retinyl bromoacetate
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0.05 mM, 76% inhibition
apo-cellular retinol-binding protein type 1
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bipyridine
CHAPS
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the addition of 0.5% (w/v) CHAPS to the assay system almost completely abolishes the 11-cis-retinol formation
FATP1c
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cytosolic C-terminal sequence from the fatty acid transport protein 1 (FATP1or SLC27A1, solute carrier family 27member1), interacts dose-dependently with the native RPE65 and markedly inhibits 11-cis retinol production by acting on the production of all-trans retinyl esters and the isomerase activity of RPE65
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N-(12-chloro-11-oxododecyl)acetamide
n-benzylidene-tert-butylamine
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14% inhibition at 0.2 mM
N-tert-butyl-alpha-phenylnitrone
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strong uncompetitive inhibition (52%) of isomerohydrolase activity in vitro at 0.2 mM
NEM
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inhibits all-trans retinyl-palmitate hydrolase and 11-cis-retinyl-palmitate hydrolase activity of the liver enzyme, inhibits all-trans-retinyl-palmitate hydrolase activity of the pigment epithelium, 11-cis-retinyl-palmitate hydrolase activity of the pigment epithelium is unaffected
nitrosobenzene
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strong inhibition of isomerohydrolase activity in vitro at 0.2 mM
PMSF
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triolein
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inhibits all-trans retinyl-palmitate hydrolase and 11-cis-retinyl-palmitate hydrolase activity of the liver enzyme, inhibits all-trans-retinyl-palmitate hydrolase activity of the pigment epithelium, 11-cis-retinyl-palmitate hydrolase activity of the pigment epithelium is unaffected
additional information
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
MYO7A protein
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the Usher 1B protein MYO7A is required for normal localization and function of the visual retinoid cycle enzyme RPE65. RPE65 normally undergoes a light-dependent translocation to become more concentrated in the central region of the RPE cells. This translocation requires MYO7A
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additional information
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0036 - 0.143
all-trans-retinyl palmitate
0.0026
an all-trans retinyl ester
pH and temperature not specified in the publication
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0.00104
an all-trans-retinyl ester
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pH and temperature not specified in the publication
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.000145
all-trans-retinyl palmitate
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purified enzyme, in 10 mM BTP buffer, pH 8.0, 100 mM NaCl, at 37°C
0.00044
an all-trans retinyl ester
pH and temperature not specified in the publication
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Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.061
N-tert-butyl-alpha-phenylnitrone
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pH and temperature not specified in the publication
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.2
1,10-phenanthroline
Bos taurus
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in 10 mM 1,3-bis[tris(hydroxymethyl)methylamino]propane, pH 8.0, 100 mM NaCl, at 37°C
0.5
2,2'-bipyridine
Bos taurus
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in 10 mM 1,3-bis[tris(hydroxymethyl)methylamino]propane, pH 8.0, 100 mM NaCl, at 37°C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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activity is 2.2fold higher in light- versus dark-adapted RPE homogenates from wild-type mice
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
8 - 9
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pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 9
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pH 5.0: 38.7% of maximal activity, pH 9.0: 46.1% of maximal activity
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
13cIMH expression is detected in the periventricular grey zone of the optic tectum and torus longitudinalis, at the fasciculus longitudinalis medialis in the medulla oblongata, and at the periventricular pretectum
Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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malignant melanoma cells are able to esterify all-trans retinol and subsequently isomerize all-trans retinyl esters into 11-cis retinol, whereas melanocytes are not able to catalyze these reactions
Manually annotated by BRENDA team
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
additional information
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in the dark, RPE65 is distributed more extensively throughout the cell, but upon exposure to light (about 100 lux, 2 h), it becomes concentrated more in the central region
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Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
60927
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x * 60927, calculated from amino acid sequence
65000
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x * 65000, SDS-PAGE
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
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x-ray crystallography
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
no modification
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Rpe65 is not palmitoylated nor does it appear to undergo other post-translational modifications at significant stoichiometry
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
hanging drop vapor diffusion method
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hanging drop vapor diffusion method, using 100 mM MES, pH 6.0, 30% (w/v) polyethylene glycol 200, and 2 mM dithiothreitol, at 22°C
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GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
0.1% (w/v) CHAPS is optimal for solubilizing RPE65 while preserving its enzymatic activity
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RPE65 is a relatively stable protein with a half-life longer than 10 h in the presence of cycloheximide
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wild type RPE65 has a half-life of more than 10 h after the addition of cycloheximide
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wild type RPE65 is a fairly stable protein, with an apparent half-life longer than 10 h, when expressed in 293A cells
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
DEAE-Sepharose column chromatography
DEAE-Sepharose column chromatography and phenyl-Sepharose CL-4B column chromatography
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Ni-NTA agarose column chromatography, gel filtration
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Ni-NTA column chromatography
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Ni-NTA resin column chromatography
purified from native bovine RPE microsomes in the presence of octyltetraoxyethylene. Triton X-100 and CHAPS are the most effective agents for extracting RPE65 from microsomal membranes
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
DNA and amino acid seuence determination and analysis, phylogenetic analyis and tree, and sequence comparisons, expression in HEK293-F cells
expressed in 293-F cells
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expressed in 293A cells; expressed in 293A cells
expressed in 293A-lecithin retinol acyltransferase cells
expressed in 293A-LRAT cells
expressed in baculovirus-infected Sf9 cells
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expressed in Escherichia coli and in 293A-LRAT cells
expressed in HEK-293-LRAT cells
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expressed in HEK-293T cells
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expressed in in baculovirus-infected Sf9 cells and in several 293T cell lines
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expressed in QBI-293A cells
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expressed in Sf9 cells and in 29T cells
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expression of wild-type enzyme and mutants in HEK-293A cells
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recombinant expression in HEK-293A cells
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recombinant expression of wild-type and mutant enzymes in HEK-293A-LRAT cells
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
2 h exposure to light effects a 25% increase over dark-adapted RPE65 levels in wild type retinas
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Myo7a-mutant mice have lower levels of RPE65
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C330Y
-
inactive
Y368H
-
inactive
F103L
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site-directed mutagenesis, the mutation reverses the enzyme isomerization product specificity from formation of 11-cis-retinol to 13-cis-retinol, product of EC 3.1.1.90. Formation of 95.7% 13-cis-retinol and 4.3% 11-cis-retinol
K222M
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site-directed mutagenesis, the mutation reverses the enzyme isomerization product specificity from formation of 11-cis-retinol to 13-cis-retinol, product of EC 3.1.1.90. Formation of 55.1% 13-cis-retinol and 44.9% 11-cis-retinol
L133S
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site-directed mutagenesis, the mutation reverses the enzyme isomerization product specificity from formation of 11-cis-retinol to 13-cis-retinol, product of EC 3.1.1.90. Formation of 71-3% 13-cis-retinol and 28.7% 11-cis-retinol
Y58N
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site-directed mutagenesis, the mutation completely reverses the enzyme isomerization product specificity from formation of 11-cis-retinol to 13-cis-retinol, product of EC 3.1.1.90. Exclusive formation of 13-cis-retinol
A132T
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50% activity compared to the wild type enzyme
A434V
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55% activity compared to the wild type enzyme
C106A
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the mutation does not affect the palmitylation status of the fragment 98-118, the mutant remains associated with the membrane. The mutant has an intact enzymatic activity similar to that of wild type enzyme
C106Y
-
site-directed mutagenesis, the mutant exhibits 16% reduced activity compared to the wild-type enzyme
C106Y/K297G
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site-directed mutagenesis, the mutant exhibits 26% reduced activity compared to the wild-type enzyme
C112A
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the mutation abolishes palmitylation in fragment 98-118 and dissociates RPE65 from the membrane. The mutant has no enzymatic activity
C231S
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55% activity compared to the wild type enzyme
C329S
-
26% activity compared to the wild type enzyme
C329S/C330S
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less than 2% activity compared to the wild type enzyme
C330Y
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less than 2% activity compared to the wild type enzyme
E417A
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less than 2% activity compared to the wild type enzyme
E417D
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the mutation alters the sub-cellular localization of the protein from membrane to the cytosol. The mutant show 5fold reduced activity compared to the wild type enzyme; the mutation decreases the stability of RPE65 and alters its sub-cellular localization, the mutant retains partial enzymatic activity (the amount of 11-cis retinol generated by E417D is approximately 5fold lower than the amount generated by wild type RPE65)
F61L
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the mutant shows 1.02% activity compared to the wild type enzyme
F61W
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the mutant shows 1.28% activity compared to the wild type enzyme
F61Y
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the mutant shows 7.57% activity compared to the wild type enzyme
G40S
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less than 2% activity compared to the wild type enzyme
G528V
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less than 2% activity compared to the wild type enzyme
H182A
-
10% activity compared to the wild type enzyme
H182Y
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10% activity compared to the wild type enzyme
H68A
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less than 2% activity compared to the wild type enzyme
H68Y
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less than 2% activity compared to the wild type enzyme
I220M
-
site-directed mutagenesis, the mutant exhibits 38% reduced activity compared to the wild-type enzyme
K294T
-
16% activity compared to the wild type enzyme
K297G
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site-directed mutagenesis, the mutant exhibits 1.7fold increased activity compared to the wild-type enzyme
K297G/L510M
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site-directed mutagenesis, the mutant exhibits 45% increased activity compared to the wild-type enzyme
K297G/N302I
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site-directed mutagenesis, the mutant exhibits 7.7% increased activity compared to the wild-type enzyme
K297G/S533A
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site-directed mutagenesis, the mutant exhibits 18% increased activity compared to the wild-type enzyme
L510M
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site-directed mutagenesis, the mutant exhibits 29% reduced activity compared to the wild-type enzyme
N170K
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site-directed mutagenesis, the mutant exhibits 1.6fold increased activity compared to the wild-type enzyme
N170K/C330T
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site-directed mutagenesis, the mutant exhibits 15% reduced activity compared to the wild-type enzyme
N170K/I220M
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site-directed mutagenesis, the mutant exhibits 69% reduced activity compared to the wild-type enzyme
N170K/K297G
-
site-directed mutagenesis, the mutant exhibits 67% increased activity compared to the wild-type enzyme
N170K/Q497P
-
site-directed mutagenesis, the mutant exhibits 16% increased activity compared to the wild-type enzyme
N302I
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site-directed mutagenesis, the mutant exhibits 25% increased activity compared to the wild-type enzyme
P363T
-
the mutation is identified in patients with Leber's congenital amaurosis, significantly decreases the stability of the enzyme, alters subcellular localization mainly to the plasma membrane and abolishes its isomerohydrolase activity
Q497P
-
site-directed mutagenesis, the mutant exhibits 26% reduced activity compared to the wild-type enzyme
R44Q
-
less than 2% activity compared to the wild type enzyme
R91W
-
the mutation from patients with retinal dystrophies decreases the stability of RPE65 protein and abolishes its isomerohydrolase activity. The mutant shows significantly decreased protein levels but unchanged mRNA levels and has a half-life of less than 2 h when expressed in 293A cells compared with the wild type
S533A
-
site-directed mutagenesis, the mutant exhibits 23% increased activity compared to the wild-type enzyme
T147A
-
the mutant shows 12.63% activity compared to the wild type enzyme
T147C
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the mutant shows 20.18% activity compared to the wild type enzyme
T147G
-
the mutant shows 30.02% activity compared to the wild type enzyme
T147S
-
the mutant shows 84% activity compared to the wild type enzyme
T147V
-
inactive
T147W
-
inactive
T147Y
-
inactive
T39R
-
site-directed mutagenesis, the mutant exhibits 22% increased activity compared to the wild-type enzyme
T39R/N170K
-
site-directed mutagenesis, the mutant exhibits 39% increased activity compared to the wild-type enzyme
T39R/N170K/C330T
-
site-directed mutagenesis, the mutant exhibits 8.4% increased activity compared to the wild-type enzyme
T39R/N170K/C330T/Q497P
-
site-directed mutagenesis, the mutant exhibits 23% reduced activity compared to the wild-type enzyme
T39R/N170K/I220M
-
site-directed mutagenesis, the mutant exhibits 10% increased activity compared to the wild-type enzyme
T39R/N170K/I220M/Q497P
-
site-directed mutagenesis, the mutant exhibits 8.6% increased activity compared to the wild-type enzyme
T39R/N170K/Q497P
-
site-directed mutagenesis, the mutant exhibits 11% increased activity compared to the wild-type enzyme
W331F
-
the mutant shows 8.34% activity compared to the wild type enzyme
W331L
-
inactive
W331Q
-
inactive
W331Y
-
the mutant shows 26.72% activity compared to the wild type enzyme
Y144D
-
the mutation is identified in patients with Leber's congenital amaurosis, significantly decreases the stability of the enzyme, alters subcellular localization mainly to the plasma membrane and abolishes its isomerohydrolase activity
Y239C
-
inactive
Y239D
-
inactive
Y239F
-
inactive
Y239L
-
inactive
Y239S
-
inactive
Y239T
-
inactive
Y239W
-
inactive
Y275F
-
the mutant shows 14.44% activity compared to the wild type enzyme
Y275I
-
inactive
Y275W
-
inactive
Y368F
-
56% activity compared to the wild type enzyme
additional information