Information on EC 2.3.1.17 - aspartate N-acetyltransferase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
2.3.1.17
-
RECOMMENDED NAME
GeneOntology No.
aspartate N-acetyltransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
acetyl-CoA + L-aspartate = CoA + N-acetyl-L-aspartate
show the reaction diagram
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-
-
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Acyl group transfer
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Alanine, aspartate and glutamate metabolism
-
-
Metabolic pathways
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-
SYSTEMATIC NAME
IUBMB Comments
acetyl-CoA:L-aspartate N-acetyltransferase
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CAS REGISTRY NUMBER
COMMENTARY hide
9029-99-6
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
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Canavan disease is a fatal, neurological disease that is caused by an interruption in the metabolism of a critical amino acid, N-acetyl-L-aspartic acid. Defects at multiple locations in the aspA gene that codes for aspartoacylase, EC 3.5.1.15, lead to mutant forms of this enzyme that are either not expressed or rapidly degraded, or have significantly impaired catalytic activity, resulting in N-acetyl-L-aspartic acid accumulation. A second gene knock-out in the Nat8l gene which codes for aspartate N-acetyltransferase, the enzyme that synthesizes N-acetyl-L-aspartic acid, reverses these adverse effects, leading to normal myelination and a decrease in Canavan disease symptoms
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
acetyl-CoA + 2,3-diaminosuccinate
CoA + ?
show the reaction diagram
-
-
-
-
?
acetyl-CoA + 3-methyl-L-aspartate
CoA + N-acetyl-3-methyl-L-aspartate
show the reaction diagram
-
-
-
-
?
acetyl-CoA + glutamate
CoA + N-acetylglutamate
show the reaction diagram
-
[14C]glutamate is acetylated with an approximately 50-fold lower affinity and a similar Vmax compared with aspartate
-
-
?
acetyl-CoA + L-aspartate
CoA + N-acetyl-L-aspartate
show the reaction diagram
-
-
-
-
?
acetyl-CoA + L-aspartate
CoA + N-acetyl-L-aspartic acid
show the reaction diagram
acetyl-CoA + L-glutamate
CoA + N-acetyl-L-glutamate
show the reaction diagram
-
reaction of EC 2.3.1.1
-
-
?
acetyl-CoA + L-glutamate
CoA + N-acetyl-L-glutamic 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
acetyl-CoA + L-aspartate
CoA + N-acetyl-L-aspartate
show the reaction diagram
-
-
-
-
?
acetyl-CoA + L-aspartate
CoA + N-acetyl-L-aspartic acid
show the reaction diagram
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
KBr
-
stimulates
KCl
-
2fold enzyme activity at 0.05 - 0.1 M
KClO3
-
stimulates
KCNS
-
stimulates
KI
-
stimulates
KNO3
-
stimulates
LiCl
-
stimulates
NaCl
-
stimulates
NaNO2
-
stimulates
NH4Cl
-
stimulates
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Acyl-AMP derivatives
-
e.g. acetyl-AMP, butyryl-AMP, strong
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benzoyl-CoA
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weak
CHAPS
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40% of activity at 12 mM, 10% of activity at 20 mM
cholamido propane sulfonate
-
-
-
CoA
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IC50: 0.42 mM
cyclohexylmaltoside
-
cymal5, high inhibition at CMC concentration
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decylmaltoside
-
-
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DMSO
-
20% inhibition at 40% v/v
dodecyl octaglycol
-
-
dodecylmaltoside
-
-
Fluoroacetyl-CoA
-
weak
glutamate
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Glutamate is a competitive inhibitor. No inhibition is observed with other amino acids, indicating that the enzyme is specific.
lauryldimethylamine-N-oxide
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complete inhibition at CMC concentration
N-acetyl-L-aspartate
-
-
N-acetyl-L-aspartic acid
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IC50: 0.85 mM
n-decyl-N,N-dimethylamine-N-oxide
-
high inhibition at CMC concentration
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N-methyl-N-nonanoyl-beta-D-glucosylamine
-
Mega-9, high inhibition at CMC concentration
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n-nonly-beta-D-glucopyranoside
-
high inhibition at CMC concentration
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NaCl
-
above 150 mM
octyl pentaglycol
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high inhibition at CMC concentration
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octyl tetraglycol
-
high inhibition at CMC concentration
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octylglucoside
-
high inhibition at CMC concentration
polymalic anhydride C10
-
-
-
polymalic anhydride C12
-
-
-
polymalic anhydride C16
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high inhibition at CMC concentration
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polymalic anhydride C4
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high inhibition at CMC concentration
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polymalic anhydride C6
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complete inhibition at CMC concentration
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polymalic anhydride C8
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-
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propionyl-CoA
-
weak
SDS
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complete inhibition at CMC concentration
sodium dodecanoyl sarcosine
-
-
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Triton X-100
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Tween 20
-
-
additional information
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effect of different detergents on the enzyme activity: non-ionic detergents such as Triton X-100 are less disruptive to protein structures than ionic detergents such as SDS, detergents such as C12E8, Tween 20 and several maltosides caused minimal disruption of the enzyme, with greater than 50% residual activity after incubation with CMC levels of each of these detergents. In contrast, significant loss of activity is observed upon incubation with C8 detergents, cymal5, octylglucoside and some shorter chain polymaleic anhydride (pmal) detergents. Ionic detergent SDS and a zwitterionic detergent lauryldimethylamine-N-oxide cause nearly complete loss of catalytic activity
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Methamphetamine
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maximum increase in activity in SH-SY5Y cells is found at 1 microM methamphetamine at 24 h, increase in activity is about 2fold
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.92
2,3-diaminosuccinate
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pH 7.4, temperature not specified in the publication
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0.36
3-methyl-L-aspartate
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pH 7.4, temperature not specified in the publication
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0.001 - 1.06
acetyl-CoA
5
glutamate
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0.09 - 3.37
L-aspartate
8.6
L-glutamate
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pH 7.4, temperature not specified in the publication
additional information
additional information
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.035
2,3-diaminosuccinate
Homo sapiens
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pH 7.4, temperature not specified in the publication
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0.0018
3-methyl-L-aspartate
Homo sapiens
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pH 7.4, temperature not specified in the publication
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0.0071
acetyl-CoA
0.023
L-glutamate
Homo sapiens
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pH 7.4, temperature not specified in the publication
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
6.1
glutamate
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-
0.56
N-acetyl-L-aspartate
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-
additional information
additional information
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Ki values for various inhibitors
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IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.42
CoA
Rattus norvegicus
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IC50: 0.42 mM
0.85
N-acetyl-L-aspartic acid
Rattus norvegicus
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IC50: 0.85 mM
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.0000063
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brain
0.000011
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spinal cord
0.0015
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activity of cloned mouse NAT8L fusion protein with N-terminal His6 tag
0.0018
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activity of cloned mouse NAT8L fusion protein with C-terminal His6 tag
0.0025
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activity of cloned wild type mouse NAT8L protein
70
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recombinant enzyme, pH 7.4, temperature not specified in the publication
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.4
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assay at
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.2 - 8
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about 13% of maximal activity at pH 5.2, about 50% of maximal activity at pH 8.0
6 - 9.5
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the enzymatic activity decreases at pH values below pH 7.5. A fit of the Vmax/Km data to a model which assumes that the protonation of a single group leads to loss of activity results in a pK value of 6.8 for a group that must be ionized for the enzyme to remain catalytically active. By contrast, the Vmax profile does not show substantial changes across the pH range
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
-
enzyme is present in both cytoplasm and mitochondria
Manually annotated by BRENDA team
-
most likely endoplasmic reticulum
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
additional information
-
not in mitochondria
-
Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
670000
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
multimer
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at least 10 bands on SDS-PAGE indicating a large enzyme complex
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
molecular modeling of the active site shows that only the amino acid aspartate, but not glutamate, can fit into the active site pocket
-
purified recombinant His-tagged truncated enzyme, X-ray diffraction structure determination and analysis
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molecular modeling of the active site shows that only the amino acid aspartate, but not glutamate, can fit into the active site pocket
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
100
-
no enzyme activity after boiling
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
0-5C, 8-10 mg protein/ml, stable for weeks
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
partial
recombinant functional and soluble dual His- and MBP-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and amylose affinity chromatography, MBP tag cleavage by 3C protease, method evaluation
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression HEK-293 cell
expression in HEK-293T cell
-
gene nat8l, sequence comparisons, subcloning of the genes for thioredoxin (TRX), glutathione S-transferase (GST) or maltose binding protein (MBP), followed by a linker region (21-68-amino acids) containing various cleavage site sequences, and then connected to the N-terminal of the nat8l gene, without the membrane anchor, recombinant functional and soluble expression of the dual affinity tagged enzyme as MBP-fusion protein in Escherichia coli strain BL21(DE3), method evaluation
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overexpressed both as native and as fusion protein with a His6 tag at the C- or the N-terminus in human embryonic kidney-293 cells expressing the large T-antigen of simian virus 40 (HEK-293T cells)
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overexpressed both as native and as fusion proteins with a His6 tag at the C- or the N-terminus in human embryonic kidney-293 cells expressing the large T-antigen of simian virus 40 (HEK-293T cells); furthermore expressed as fusion protein in rat embryonal cortical neurons and in Chinese-hamster ovary cells
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C128A
-
mutation in region 4, 94% of wild-type expression, 88% of wild-type activity
C139A
-
mutation in region 4, 70% of wild-type expression, 126% of wild-type activity
D168A
-
mutation in region 5, 46% of wild-type expression, no residual activity
D168E
-
mutation in region 5, 63% of wild-type expression, 7% of wild-type activity
E101A
-
mutation in region 3, 97% of wild-type expression, 42% of wild-type activity
E101D
-
mutation in region 3, 61% of wild-type expression, 99% of wild-type activity
P142A
-
mutation in region 4, 113% of wild-type expression, 116% of wild-type activity
R133A
-
mutation in region 4, 78% of wild-type expression, 64% of wild-type activity
R133K
-
mutation in region 4, 69% of wild-type expression, 89% of wild-type activity
R220A
-
mutation in region 5, 101% of wild-type expression, 14% of wild-type activity
R220K
-
mutation in region 5, 104% of wild-type expression, 25% of wild-type activity
R81A
-
mutation in region 3, 57% of wild-type expression, 37% of wild-type activity
R81K
-
mutation in region 3, 90% of wild-type expression, 82% of wild-type activity
S132F/R133F
-
mutation in region 4, 92% of wild-type expression, 41% of wild-type activity
additional information
-
transfection of truncated forms of isoform NAT8L into HEK-293T cells indicates that the 68 N-terminal residues, i.e. regions 1 and 2, have no importance for the catalytic activity and the subcellular localization of the enzyme. The membrane region, i.e. region 4, is necessary and sufficient to target isoform NAT8L to the endoplasmic reticulum