Information on EC 3.2.1.46 - galactosylceramidase

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The expected taxonomic range for this enzyme is: Euarchontoglires

EC NUMBER
COMMENTARY hide
3.2.1.46
-
RECOMMENDED NAME
GeneOntology No.
galactosylceramidase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
a D-galactosyl-N-acylsphingosine + H2O = D-galactose + a ceramide
show the reaction diagram
-
-
-
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of O-glycosyl bond
hyrolysis of O-glycosyl bond
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Sphingolipid metabolism
-
-
SYSTEMATIC NAME
IUBMB Comments
D-galactosyl-N-acylsphingosine galactohydrolase
cf. EC 3.2.1.62 glycosylceramidase.
CAS REGISTRY NUMBER
COMMENTARY hide
9027-89-8
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
Uniprot
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
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
4-methylumbelliferyl beta-galactoside + H2O
methylumbelliferone + beta-D-galactose
show the reaction diagram
4-methylumbelliferyl-beta-D-galactopyranoside + H2O
4-methylumbelliferone + beta-D-galactopyranose
show the reaction diagram
5-bromo-3-chloro-beta-galactopyranoside + H2O
?
show the reaction diagram
-
-
-
?
6-hexadecanoylamino-4-methylbelliferyl-beta-D-galactopyranoside + H2O
6-hexadecanoylamino-4-methylbelliferone + beta-D-galactopyranose
show the reaction diagram
D-galactosyl-alkyl-acyl-glycerol + H2O
?
show the reaction diagram
-
a precursor of the seminolipid
-
-
?
D-galactosyl-N-acylsphingosine + H2O
?
show the reaction diagram
D-galactosyl-N-acylsphingosine + H2O
D-galactose + N-acylsphingosine
show the reaction diagram
D-galactosylceramide + H2O
D-galactose + ceramide
show the reaction diagram
-
-
-
-
?
D-galactosylsphingoside + H2O
D-galactose + sphingosine
show the reaction diagram
-
i.e. psychosine
-
-
?
galactocerebroside + H2O
D-galactose + N-acylceramide
show the reaction diagram
galactosylceramide + H2O
D-galactose + N-acylceramide
show the reaction diagram
GM1 ganglioside + H2O
D-galactose + N-acylceramide
show the reaction diagram
lactosylsphingosine + H2O
lactose + sphingosine
show the reaction diagram
N-stearoyl psychosine + H2O
D-galactose + N-stearoylsphingosine
show the reaction diagram
-
-
-
-
?
psychosine + H2O
D-galactose + sphingosine
show the reaction diagram
additional information
?
-
-
the enzyme is required for normal sperm maturation and function, enzyme deficiency leads to degeneration of oligodendrocytes, severe demyelination, and causes sperm abnormalities in the mouse model of human globoid cell leukodystrophy or Krabbe disease, mutant twitcher mice show reduced size of testis and sperm acrosomal membrane which is redundant, detached from the nucleus and folded over
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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
D-galactosyl-N-acylsphingosine + H2O
?
show the reaction diagram
D-galactosyl-N-acylsphingosine + H2O
D-galactose + N-acylsphingosine
show the reaction diagram
-
-
-
-
?
D-galactosylceramide + H2O
D-galactose + ceramide
show the reaction diagram
-
-
-
-
?
D-galactosylsphingoside + H2O
D-galactose + sphingosine
show the reaction diagram
-
i.e. psychosine
-
-
?
additional information
?
-
-
the enzyme is required for normal sperm maturation and function, enzyme deficiency leads to degeneration of oligodendrocytes, severe demyelination, and causes sperm abnormalities in the mouse model of human globoid cell leukodystrophy or Krabbe disease, mutant twitcher mice show reduced size of testis and sperm acrosomal membrane which is redundant, detached from the nucleus and folded over
-
-
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Cl-
-
activating
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2-hydroxydecanoyl DL-erythro-3-phenyl-2-amino-1,3-propanediol
-
2-hydroxy fatty acid amides of phenylaminopropanediol, noncompetitive
2-hydroxydodecanoyl DL-erythro-3-phenyl-2-amino-1,3-propanediol
-
2-hydroxy fatty acid amide of phenylaminopropanediol, noncompetitive
2-hydroxyhexadecanoyl DL-erythro-3-phenyl-2-amino-1,3-propanediol
-
2-hydroxy fatty acid amides of phenylaminopropanediol, noncompetitive
2-hydroxyoctadecanoyl DL-erythro-3-phenyl-2-amino-1,3-propanediol
-
2-hydroxy fatty acid amides of phenylaminopropanediol, noncompetitive
2-hydroxytetradecanoyl DL-erythro-3-phenyl-2-amino-1,3-propanediol
-
2-hydroxy fatty acid amides of phenylaminopropanediol, noncompetitive
6-hexadecanoylamino-4-methylbelliferyl-beta-D-galactopyranoside
D-galactose
Galactonolactone
-
-
galactonyl hydrazide
-
-
-
lactose
lactosyl ceramide
-
-
N-(6-aminohexyl)-D-galactoside
-
-
-
N-decanoyl psychosine
-
competitve and noncompetitive
N-dodecanoyl psychosine
-
competitve and noncompetitive
N-ethyl psychosine
-
competitve and noncompetitive
N-hexyl psychosine
-
competitve and noncompetitive
N-octanoyl psychosine
-
competitve and noncompetitive
taurocholate
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Activator protein
-
purified from normal spleen
-
cholate
-
-
phosphatidylcholine
-
from 0.02 mg to 0.1 mg
phosphatidylinositol
-
from 0.02 mg to 0.05 mg
phosphatidylserine
taurocholate
Triton X-100
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.47
4-methylumbelliferyl-beta-D-galactopyranoside
0.017 - 0.15
6-hexadecanoylamino-4-methylbelliferyl-beta-D-galactopyranoside
0.005 - 0.025
galactocerebroside
0.02 - 0.2
galactosylceramide
0.11 - 0.67
galactosylsphingosine
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.000008
-
GALC enzyme activity in the posterior part of the brain of single intracerebroventricular-GALC injected mice
0.0000087
-
GALC enzyme activity in the anterior part of the brain of single intracerebroventricular-GALC injected mice
0.12
-
-
70
-
with plasmid encoding GALC-TMH transfected 293T cell, GALC units per mg of cell extract
120
-
with plasmid encoding GALC-MH transfected 293T cell, GALC units per mg of cell extract
130
-
with plasmid encoding unmodified GALC transfected 293T cell, GALC units per mg of cell extract
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3.75
-
without taurocholate
4.6
-
galactocerebroside, cholate system
4.7
-
phosphatidylserine activated
5.2
-
6-hexadecanoylamino-4-methylbelliferyl-beta-D-galactopyranoside, cholate system
5.4
-
with taurocholate
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
2.5 - 5.5
3.5 - 5.3
-
pH 3.5: about 10% of maximal activity, pH 5.3: about 40% of maximal activity
3.5 - 6
-
-
3.5 - 5.3
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pH 3.5: about 10% of maximal activity, pH 5.3: about 40% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
-
level of activity in umbilical cord blood is comparable ro that in adult blood
Manually annotated by BRENDA team
-
hippocampal pyramidal neuron, cerebellar neuron
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30000
-
gel filtration, SDS-PAGE, N-terminal sequencing
50000
-
urine, gel filtration, SDS-PAGE, N-terminal sequencing
70000
-
gel filtration, SDS-PAGE, N-terminal sequencing
77000
-
amino acid analysis
90000
-
gel filtration, SDS-PAGE, N-terminal sequencing
121000
-
gel filtration, monomer
640000
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gel filtration chromatography
750000
-
gel filtration, SDS-PAGE, mutant and normal protein
760000
-
gel filtration, hexamer
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
-
2 * 125000, aggregation from monomer to dimer, from dimer to tetramer or hexamer, SDS-PAGE
hexamer
-
6 * 125000, active form in vivo, SDS-PAGE
monomer
-
1 * 125000, aggregation from monomer to dimer, from dimer to tetramer or hexamer, SDS-PAGE
tetramer
-
4 * 125000, aggregation from monomer to dimer, from dimer to tetramer or hexamer, SDS-PAGE
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycoprotein
-
-
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
crystal structures of GALC and the GALC-D-galactose product complex, to 2.1 and 2.4 A resolution, respectively. The overall fold comprises a central triosephosphate isomerase barrel, a beta-sandwich domain, and a lectin domain. The overall fold of GALC is unchanged upon galactose binding, the core of the binding pocket being formed by the long loops on the C-terminal face of the TIM barrel. Loops from both the beta-sandwich and lectin domains also contribute to the substrate-binding pocket, and mutations involved in Krabbe's disease are widely distributed throughout the protein
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
52
-
35 min, 50% loss of activity, normal enzyme; 4 min, 50% loss of activity, mutant enzyme
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
bovine serum albumin for stabilization
-
urea, 50% loss of activity of the mutant enzyme at 1.3 M urea, 50% loss of activity of the normal enzyme at 5.6 M urea
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
partially
to homogeneity, Krabbe disease
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression in CHO cells
-
expression in lentiviral GALC vectors
-
expression vectors for different fusion-proteins are constructed: GALC, unmodified enzyme, GALC-MH: fusion protein containing a C-terminal myc-tag and six His residues, GALC-TMH: fusion protein containing in addition a Tat-PTD, the protein transduction domain derived from the HIV-1 Tat protein
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exression in lentiviral GALC vectors
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mutation in Krabbe disease
nonsense mutation in Krabbe disease
-
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
E114K
residue in TIM barrel, mutation is likely to result in severe misfolding, crystallization data
E215K
residue is exposed on the surface of the TIM barrel, mutation confers an opposite charge on the same face as the substrate-binding pocket, crystallization data
G537R
resiude in lectin domain, mutation is likely to result in severe misfolding, crystallization data
GALCdelta-MH
-
mutant containing a C-terminal myc-tag and six His residues with the last 11 amino acids at the C-terminus of GALC deleted
L364R
resiude in beta-sandwich, mutation is likely to result in severe misfolding, crystallization data
L629R
resiude in lectin domain, mutation is likely to result in severe misfolding, crystallization data
S257F
resiude in TIM barrel, mutation is likely to result in severe misfolding, crystallization data
W410G
resiude in beta-sandwich, mutation is likely to result in severe misfolding, crystallization data
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
medicine