Information on EC 6.1.1.12 - aspartate-tRNA ligase

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

EC NUMBER
COMMENTARY hide
6.1.1.12
-
RECOMMENDED NAME
GeneOntology No.
aspartate-tRNA ligase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + L-aspartate + tRNAAsp = AMP + diphosphate + L-aspartyl-tRNAAsp
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
aminoacyl group transfer
Aminoacylation
esterification
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
tRNA charging
-
-
aspartate and asparagine metabolism
-
-
Aminoacyl-tRNA biosynthesis
-
-
SYSTEMATIC NAME
IUBMB Comments
L-aspartate:tRNAAsp ligase (AMP-forming)
-
CAS REGISTRY NUMBER
COMMENTARY hide
9027-32-1
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
C4LZN0
UniProt
Manually annotated by BRENDA team
Escherichia coli B / ATCC 11303
-
-
-
Manually annotated by BRENDA team
gene aspS
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
overexpressed in Escherichia coli
-
-
Manually annotated by BRENDA team
overexpressed in Escherichia coli
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
Thermus thermophilus HB8 / ATCC 27634 / DSM 579
-
SwissProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
LBSL, i.e. leukoencephalopathy with brain stem and spinal cord involvement and lactate elevation, is a monogenic disease associated with a large variety of mutations affecting the human nuclear gene DARS2, encoding mt-AspRS, overview
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
adenosine 5’-[beta,gamma-imido]-triphosphate + L-Asp
AMP-Asp + imidodiphosphate
show the reaction diagram
AMP-Asp + diphosphate
ATP + L-Asp
show the reaction diagram
ATP + Asp + tRNAAsn
AMP + diphosphate + aspartyl-tRNAAsn
show the reaction diagram
-
discriminating AspRS gains the ability to form Asp-tRNAAsn in vitro when the W26H or K85P changes are introduced independently or in combination
-
-
?
ATP + Asp + tRNAAsp
AMP + diphosphate + aspartyl-tRNAAsp
show the reaction diagram
-
-
-
-
?
ATP + D-aspartate + tRNAAsp
AMP + diphosphate + D-aspartyl-tRNAAsp
show the reaction diagram
-
aspartyl-tRNA synthetase can misacylate tRNAAsp with D-aspartate instead of its usual substrate, L-Asp, substrate specificity and molecular dynamics simulations, overview
-
-
?
ATP + L-Asp + tRNAAsp
AMP + diphosphate + aspartyl-tRNAAsp
show the reaction diagram
-
low reaction with mutant tRNAAsp with A instead of G at position 73 (tRNAAspA73)
-
-
?
ATP + L-asparagine + tRNAAsp
AMP + diphosphate + L-asparaginyl-tRNAAsp
show the reaction diagram
-
-
-
?
ATP + L-aspartate + tRNAAsn
AMP + diphosphate + L-aspartyl-tRNAAsn
show the reaction diagram
ATP + L-aspartate + tRNAAsn
AMP + diphosphate + L-aspartyl-tRNAAsp
show the reaction diagram
ATP + L-aspartate + tRNAAsp
AMP + diphosphate + aspartyl-tRNAAsp
show the reaction diagram
ATP + L-aspartate + tRNAAsp
AMP + diphosphate + L-aspartyl-tRNAAsp
show the reaction diagram
GTP + L-aspartate + tRNAAsp
GMP + diphosphate + L-aspartyl-tRNAAsp
show the reaction diagram
UTP + L-aspartate + tRNAAsp
UMP + diphosphate + L-aspartyl-tRNAAsp
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
ATP + L-aspartate + tRNAAsn
AMP + diphosphate + L-aspartyl-tRNAAsn
show the reaction diagram
ATP + L-aspartate + tRNAAsp
AMP + diphosphate + aspartyl-tRNAAsp
show the reaction diagram
ATP + L-aspartate + tRNAAsp
AMP + diphosphate + L-aspartyl-tRNAAsp
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Co2+
-
can replace Mg2+, with 12% efficiency in activation of ATP-diphosphate exchange, with 34% efficiency in aminoacylation
K2HPO4
-
enzyme form PC-3 is stimulated, optimal concentration 75 mM, enzyme form PC-1 and enzyme form PC-2 are inhibited
KCl
-
enzyme form PC-3 is stimulated, optimal concentration 75 mM, enzyme form PC-1 and enzyme form PC-2 are inhibited
Mn2+
-
can replace Mg2+, with 35% efficiency in activation of ATP-diphosphate exchange, with 56% efficiency in aminoacylation
PO43-
-
appears to act synergistically with K+ in stimulation of enzyme form PC-3, no effect on either enzyme form PC-1 or enzyme form PC-2
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
(3S)-3-amino-5-((((2R,3S,4R,5R)-5-(6-amino-9H-purin-9-yl)-3,4-dihydroxytetrahydrofuran-2-yl)oxy)(hydroxy)phosphoryl)-4-oxopentanoic acid
-
-
1-[(5E)-5-[(2,4-dichlorophenyl)methylidene]-4-oxo-4,5-dihydro-1,3-thiazol-2-yl]piperidine-4-carboxamide
84.3% residual activity at 0.3 mM
2-aminomalonic acid
-
inhibits transfer of Asp to tRNA
5'-O-[N-(L-aspartyl)sulfamoyl]adenosine
CaCl2
-
1 mM, more than 50% inhibition
CH3COO-
-
2.0 M, 50% inhibition of ATP-diphosphate exchange
ClO3-
-
0.1 M, 50% inhibition of ATP-diphosphate exchange
CNS-
-
0.1 M, 50% inhibition of ATP-diphosphate exchange
Cs+
-
1.1 M, 50% inhibition of ATP-diphosphate exchange, 0.16 M, 50% inhibition of aminoacylation
diethyldicarbonate
-
reversed by hydroxylamine
erythro-3-hydroxyaspartic acid
-
inhibits transfer of Asp to tRNA
ethyl 1-[(5E)-5-[(4-chlorophenyl)methylidene]-4-oxo-4,5-dihydro-1,3-thiazol-2-yl]piperidine-4-carboxylate
87.5% residual activity at 0.1 mM
F-
-
-
gamma-aminobutyric acid
-
-
heptapeptide-nucleotide microcin C
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i.e. McC, upon its entry into a susceptible cell, McC is processed to release a nonhydrolyzable aspartyl-adenylate that inhibits aspartyl-tRNA synthetase, leading to the cessation of translation and cell growth. The rate-limiting step of McC processing in vitro is deformylation of the first methionine residue of McC
IO3-
-
0.04 M, 50% inhibition of ATP-diphosphate exchange
K+
-
2.3 M, 50% inhibition of ATP-diphosphate exchange, 0.22 M, 50% inhibition of aminoacylation
K2HPO4
-
inhibits enzyme forms PC-1 and PC-2, PC-3 is stimulated (optimal concentration 75 mM)
KCl
-
inhibits enzyme forms PC-1 and PC-2, PC-3 is stimulated (optimal concentration 75 mM)
L-aspartol adenylate
-
-
L-aspartol-adenylate
L-aspartyl adenylate
-
-
Li+
-
0.6 M, 50% inhibition of ATP-diphosphate exchange, 0.11 M, 50% inhibition of aminoacylation
Microcin C
-
a ribosome-synthesized heptapeptide that contains a modified adenosine monophosphate covalently attached to the C-terminal aspartate, a potent inhibitor of bacterial growth, that targets the translation process via its degradation product, a modified aspartyl-adenylate containing an N-acylphosphoramidate linkage, overview, Microcystin is recombinantly expressed in Escherichia coli strain TG1
microcin C7-C51
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an antimicrobial nucleotide peptide that targets aspartyl-tRNA synthetase and inhibits translation. Fragmentation and analysis of fragment structure and inhibitory potency, overview
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Na+
-
1.4 M, 50% inhibition of ATP-diphosphate exchange, 0.18 M , 50% inhibition of aminoacylation
NaCl
-
inhibits enzyme forms PC-1 and PC-2 much more than PC-3
NH4+
-
0.9 M, 50% inhibition of ATP-diphosphate exchange, 0.14 M, 50% inhibition of aminoacylation
NiCl2
-
0.1 mM more than 50% inhibition
NO3-
-
0.7 M, 50% inhibition of ATP-diphosphate exchange
p-chloromercuribenzoate
-
-
succinate
-
50% inhibition at 210 mM
threo-3-hydroxyaspartic acid
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inhibits transfer of Asp to tRNA
threo-3-Methylaspartic acid
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inhibits transfer of Asp to tRNA
tRNA
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above 0.4 mg/ml
additional information
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the yeast AspRS initiates retro-inhibition of its expression, overview
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
putrescine
spermidine
spermine
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
8
2-aminomalonic acid
-
ATP-diphosphate exchange
0.003 - 4.6
Asp
1.2 - 1.4
aspartate
0.03
aspartic acid
pH 7.2, 70°C
0.023 - 1.11
ATP
0.32
L-Asp
-
-
0.0015 - 0.21
L-aspartate
0.005
L-aspartic acid
pH 7.2, 70°C
24
threo-3-hydroxyaspartic acid
-
ATP-diphosphate exchange
0.000063 - 0.0091
tRNAAsn
0.000013 - 0.08
tRNAAsp
1.3
tRNAAspA73
-
pH 7.8, 28°C
-
additional information
additional information
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2 - 18
Asp
16.9
aspartyl-tRNA
-
-
5.9 - 8.5
ATP
0.0008 - 0.12
tRNAAsn
0.00016 - 79
tRNAAsp
0.0029
tRNAAspA73
-
pH 7.8, 28°C
-
additional information
additional information
-
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2.9 - 6
tRNAAsn
3.7 - 293
tRNAAsp
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.000123
(3S)-3-amino-5-((((2R,3S,4R,5R)-5-(6-amino-9H-purin-9-yl)-3,4-dihydroxytetrahydrofuran-2-yl)oxy)(hydroxy)phosphoryl)-4-oxopentanoic acid
-
-
0.0000098 - 0.00039
5'-O-[N-(L-aspartyl)sulfamoyl]adenosine
1
CaCl2
-
-
0.045
L-aspartol adenylate
-
pH 7.5, 37°C
0.00033 - 0.045
L-aspartol-adenylate
6000
Li+
-
ATP-diphosphate exchange
0.1
NiCl2
-
-
additional information
additional information
-
Microcystin C in vivo inhibition kinetics
-
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0499
1-[(5E)-5-[(2,4-dichlorophenyl)methylidene]-4-oxo-4,5-dihydro-1,3-thiazol-2-yl]piperidine-4-carboxamide
Mycobacterium tuberculosis;
P9WFW3
at pH 7.6 and 37°C
0.0591
ethyl 1-[(5E)-5-[(4-chlorophenyl)methylidene]-4-oxo-4,5-dihydro-1,3-thiazol-2-yl]piperidine-4-carboxylate
Mycobacterium tuberculosis;
P9WFW3
at pH 7.6 and 37°C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.00002
native enzyme, cell extract, 37°C
0.000083
native enzyme, cell extract, 37°C
0.017
-
wild-type enzyme
0.025
-
recombinant wild-type enzyme
0.0336
purified recombinant enzyme, 37°C
0.034
-
truncated enzyme
0.0405
-
enzyme form AspRS2
0.043
recombinant enzyme, Escherichia coli DH5alpha cell extract, 37°C
0.0452
-
enzyme form AspRS1
0.055
recombinant enzyme, Escherichia coli BL21 cell extract, 37°C
0.063
-
recombinant mutant D560V
0.097
-
recombinant mutant R263Q
0.11
-
65°C, pH is not specified in the publication; 65°C, pH not specified in the publication
0.12
-
recombinant mutant L626V
0.124
recombinant enzyme, Escherichia coli BL21 cell extract, 37°C
0.15
-
recombinant mutant Q184K
0.155
recombinant enzyme, Escherichia coli DH5alpha cell extract, 37°C
0.321
purified recombinant enzyme, 37°C
1.75
-
recombinant mutant C152F
3.7
-
purified recombinant His6-tagged DRS enzyme
10.9
-
purified recombinant His6-tagged DRS-ubiquitin fusion enzyme
11.5
-
purified recombinant biotin-tagged DRS-ubiquitin fusion enzyme
13.5
-
purified recombinant His6-tagged DRS-small ubiquitin-like modifier fusion enzyme
17
-
purified recombinant GST-tagged DRS enzyme
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.3
assay at
7.8 - 8
-
potassium Hepes buffer
8.5
-
Tris/cacodylate buffer
additional information
-
very low kinetic effiency at 17°C
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.5 - 9
-
pH 5.5: about 60% of maximal activity, pH 9.0: about 30% of maximal activity
5.5 - 8
-
pH 5.5: about 60% of maximal activity, pH 8.0: about 65% of maximal activity
7.8 - 9.1
-
7.8: sharp loss of activity below, 9.1: plateau of high activity up to pH 9.1
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37 - 50
-
assay at 37°C or 50°C
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
35 - 75
-
35°C: about 60% of maximal activity, 75°C: about 50% of maximal activity
45 - 85
-
45°C: about 80% of maximal activity, 85°C: about 25% of maximal activity
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.23
-
calculated from sequence
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
PDB
SCOP
CATH
UNIPROT
ORGANISM
C4LZN0
Entamoeba histolytica;
Escherichia coli (strain K12);
Homo sapiens;
Saccharomyces cerevisiae (strain ATCC 204508 / S288c);
Thermococcus kodakarensis (strain ATCC BAA-918 / JCM 12380 / KOD1);
Thermus thermophilus;
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
12870
-
1 * 12870, calculated from amino acid sequence
15500
-
gel filtration
48329
x * 48329, amino acid sequence determination
50893
-
x * 50893, calculated from sequence
53000
-
2 * 53000, gel electrophoresis under denaturing conditions
60800
theoretical
61000
-
2 * 61000, sedimentation equilibrium under denaturing conditions
62900
theoretical
63000
-
2 * 63000, SDS-PAGE
65912
-
x * 65912, crystal structure determination
66000
determined by SDS-PAGE
66029
x * 66029, amino acid sequence determination
66030
x * 66030, calculation from nucleotide sequence
83000 - 100000
-
sucrose density gradient centrifugation
89000 - 106000
-
sucrose density gradient centrifugation
96000
-
crystal structure determination
116000
-
sedimentation equilibrium determination, neutron scattering
117000
-
gel filtration, also a smaller peak of MW 57000 detected
119000
-
sedimentation equilibrium determination
122000
124000
-
gel filtration
132000
-
gel filtration
150000
-
gel filtration, PC-1 and PC-2
156000
-
nondenaturing PAGE
205000
-
gel filtration
260000
-
gel filtration
500000
-
gel filtration, PC-3, possibly PC-3 represents a large complex of aminoacyl-tRNA synthetases
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homodimer
monomer
additional information
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
lipoprotein
-
high molecular weight aminoacyl-tRNA synthetase complex contains lipid. Delipidation does not affect the size or activity of the complex, but a variety of functional and structural properties of individual synthetases in the complex are altered: sensitivity to salts plus detergents, temperature inactivation, hydrophobicity, sensitivity to protease digestion
no glycoprotein
-
-
additional information
-
the enzyme does not perform autoaspartylation in vivo
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
the crystal structure of aspartyl-tRNA synthetase is determined at 2.8 A resolution
C4LZN0;
analysis of the 2.6-A resolution crystal structure of Escherichia coli AspRS with bound aspartyl-adenylate, AspAMP, molecular dynamics simulations
-
of enzyme complexed with its cognate tRNA
-
purified recombinant enzyme, in ammonium sulfate, NaCl, bis Tris propane, isopropanol, X-ray diffraction structure determination at 2.7 A resolution and structure analysis
-
hanging drop vapor diffusion method, in 30% (w/v) PEG 4000, 100 mM ammonium sulfate, 100 mM sodium citrate at pH 5.6
-
hanging drop vapor diffusion method, using 7% (w/v) PEG 4000, 0.1 M ammonium sulfate, 0.1 M sodium citrate pH 5.6
vapor diffusion method, using 10% (m/v) PEG-3350 and 0.5M of ammonium sulfate in 25 mM Bis-Tris pH 5.5