Information on EC 3.4.19.5 - beta-aspartyl-peptidase

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

EC NUMBER
COMMENTARY hide
3.4.19.5
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RECOMMENDED NAME
GeneOntology No.
beta-aspartyl-peptidase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
Cleavage of a beta-linked Asp residue from the N-terminus of a polypeptide
show the reaction diagram
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of peptide bond
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CAS REGISTRY NUMBER
COMMENTARY hide
37288-74-7
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
enzyme from human faeces, present in healthy individuals, absent in antibiotic-treated patients
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-
Manually annotated by BRENDA team
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H0VQC8
UniProt
Manually annotated by BRENDA team
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UniProt
Manually annotated by BRENDA team
yellow lupin with roots infected with Bradyrhizobium sp.
UniProt
Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
the enzyme belongs to the N-terminal nucleophile hydrolase family
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
alpha-Asp-Leu + H2O
L-Asp + L-Leu
show the reaction diagram
-
-
-
-
?
Asp-Gly-Ala + H2O
Asp + Gly-Ala
show the reaction diagram
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55% of the activity with beta-aspartylglycine
-
?
Asp-Gly-Val + H2O
Asp + Gly-Val
show the reaction diagram
-
13% of the activity with beta-aspartylglycine
-
?
beta-Asp-Ala + H2O
Asp + Ala
show the reaction diagram
beta-Asp-Gln + H2O
Asp + Gln
show the reaction diagram
-
-
-
?
beta-Asp-Gly + H2O
Asp + Gly
show the reaction diagram
beta-Asp-Gly-Gly + H2O
Asp + Gly-Gly
show the reaction diagram
-
95% of the activity with beta-aspartylglycine
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?
beta-Asp-His + H2O
Asp + His
show the reaction diagram
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-
-
-
?
beta-Asp-Ile + H2O
Asp + Ile
show the reaction diagram
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37% of the activity with beta-aspartylglycine
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?
beta-Asp-Leu + H2O
Asp + Leu
show the reaction diagram
beta-Asp-Lys + H2O
Asp + Lys
show the reaction diagram
-
-
-
-
?
beta-Asp-Met + H2O
Asp + Met
show the reaction diagram
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82% of the activity with beta-aspartylglycine
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?
beta-Asp-Phe + H2O
Asp + Phe
show the reaction diagram
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-
-
-
?
beta-Asp-Phe + H2O
L-Asp + L-Phe
show the reaction diagram
H0VQC8;
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-
-
?
beta-Asp-Ser + H2O
Asp + Ser
show the reaction diagram
beta-Asp-Thr + H2O
Asp + Thr
show the reaction diagram
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29% of the activity with beta-aspartylglycine
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?
beta-Asp-Val + H2O
Asp + Val
show the reaction diagram
-
28% of the activity with beta-aspartylglycine
-
?
beta-L-Asp-L-Ala + H2O
L-Asp + L-Ala
show the reaction diagram
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-
-
?
beta-L-Asp-L-Leu + H2O
L-Asp + L-Leu
show the reaction diagram
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-
-
?
beta-L-Asp-L-Lys + H2O
L-Asp + L-Lys
show the reaction diagram
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-
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?
beta-L-Asp-L-Phe + H2O
L-Asp + L-Phe
show the reaction diagram
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-
-
?
beta-L-Asp-L-Phe methyl ester + H2O
L-Asp + L-Phe methyl ester
show the reaction diagram
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-
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?
beta-L-aspartyl-L-leucine + H2O
?
show the reaction diagram
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-
?
iso-Asp-Gly + H2O
?
show the reaction diagram
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?
iso-Asp-Leu + H2O
?
show the reaction diagram
best substrate
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-
?
L-Asp beta-methyl ester + H2O
L-Asp + methanol
show the reaction diagram
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?
L-asparagine + H2O
L-aspartate + NH3
show the reaction diagram
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?
L-aspartic acid beta-(7-amido-4-methylcoumarin) + H2O
L-aspartic acid + 7-amino-4-methylcoumarin
show the reaction diagram
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?
additional information
?
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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
L-asparagine + H2O
L-aspartate + NH3
show the reaction diagram
Q9ZSD6
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-
?
additional information
?
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Asp-PSI[PO2CH2]-LeuOH
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p-hydroxymercuribenzoate
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Zn2+
inhibits autoproteolysis, other divalent cations than Zn2+ have no effect on the process
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2-mercaptoethanol
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slight activation
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
5
alpha-Asp-Leu
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wild-type enzyme, pH 8.1, 30°C
3.7
beta-Asp-Ala
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wild-type enzyme, pH 8.1, 30°C
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18
beta-Asp-Gly
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wild-type enzyme, pH 8.1, 30°C
3.7
beta-Asp-His
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wild-type enzyme, pH 8.1, 30°C
0.09 - 34
beta-Asp-Leu
0.91
beta-Asp-Lys
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wild-type enzyme, pH 8.1, 30°C
0.23 - 0.41
beta-Asp-Phe
0.8
beta-aspartyl-L-leucine
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0.136
beta-L-aspartyl-L-leucine
pH 7.5, 22°C
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
15.7
alpha-Asp-Leu
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wild-type enzyme, pH 8.1, 30°C
213
beta-Asp-Ala
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wild-type enzyme, pH 8.1, 30°C
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0.93
beta-Asp-Gly
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wild-type enzyme, pH 8.1, 30°C
20.8
beta-Asp-His
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wild-type enzyme, pH 8.1, 30°C
0.00062 - 104
beta-Asp-Leu
58
beta-Asp-Lys
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wild-type enzyme, pH 8.1, 30°C
1.98 - 16.9
beta-Asp-Phe
0.813
beta-L-aspartyl-L-leucine
pH 7.5, 22°C
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
4.83
beta-Asp-Phe
H0VQC8;
pH 7.5, 37°C
5.98
beta-L-aspartyl-L-leucine
pH 7.5, 22°C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7 - 9
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Tris-HCl buffer
7.4
assay at
7.5
assay at
7.5 - 8
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sodium phosphate buffer
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
22
assay at room temperature
37
assay at
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
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Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
UNIPROT
ORGANISM
Escherichia coli (strain K12);
Homo sapiens;
Q9ZSD6
Lupinus luteus;
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
14550
mass spectroscopy, beta-subunit
15000
H0VQC8;
beta-subunit, SDS-PAGE
18000
Western blot, reducing conditions, detected in soluble and insoluble fractions
18500
mass spectroscopy, alpha-subunit
20000
H0VQC8;
alpha-subunit, SDS-PAGE
33020
mass spectroscopy, precursor enzyme
35000
H0VQC8;
inactive precursor, SDS-PAGE
75000
about, recombinant His-tagged enzyme, gel filtration
320000
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
heterodimer
alphabeta, 1 * 23000, alpha-subunit + 1 * 14000, beta-subunit, SDS-PAGE, 1 * 22893, His-tagged alpha-subunit + 1 * 13605, beta-subunit, mass spectrometry
tetramer
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the enzyme is dimer of heterodimers (alphabeta)2
additional information
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the quarternary structure of the enzyme is octameric and can be described as a tetramer od dimers. Each subunit folds into two distinct domains
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
proteolytic modification
H0VQC8;
inactive precursor undergoes a self-activation process, converting it from an uncleaved precursor of 35000 Da into alpha- and beta-subunits of 20000 and 15000 Da, respectively. Glycine stimulates cleavage in a dose-dependent manner
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
uncleaved (inactive) as well as the cleaved (active), aspartate-bound structures. Cleavage does not greatly alter the overall structure of the enzyme with the exception of the conformational change in the conserved HGG loop that coincides with cleavage
H0VQC8;
hanging drop method of vapor diffusion, the best crystals are observed growing at 25°C from 10% poly(ethylene glycol)8000, 100 mM homopipes, pH 5, in the presence of MgCl2
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hanging drop vapor diffusion method, 1.9 A resolution
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hanging drop vapor diffusion method. X-ray crystal structure of the D285N mutant complexed with beta-Asp-His
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hanging-drop vapour-diffusion method, crystal structure of EcAIII at 1.65 A resolution
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sitting drop vapor diffusion method, crystal structure in the absence and presence of the phosphinic inhibitor Asp-PSI[PO2CH2]-LeuOH
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
60
purified recombinant His-tagged enzyme, stable up to
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
stability is dependent on the presence of sodium ions
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stable to freezing and thawing in 0.01 M sodium phosphate buffer
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unstable to dialysis against water or 0.01 M Tris-HCl buffer, pH 8.0, if 0.01 M NaCl is included in the dialysis solution, 80% of the activity is recovered
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
room temperature, stable for 5 days
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
recombinant His-tagged enzyme in Escherichia coli strain BL21(DE3)pLysS by nickel affinity chromatography and gel filtration
using Ni-NTA chromatography. hASRGL1 purification by immobilized metal ion affinity chromatography yields 30 mg of protein/l with a purity of more than 90% as determined by SDS-PAGE
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
DNA and amino acid sequence determination and analysis, sequence comparisons and phylogenetic analysis, recombinant expression of N-terminally His-tagged enzyme in Escherichia coli strain BL21(DE3)pLysS
expressed as a His-tagged fusion protein in Escherichia coli
expression in Escherichia coli
H0VQC8;
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D285A
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kcat/Km for beta-Asp-Leu is 85000fold lower than wild-type value
D285N
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kcat/Km for beta-Asp-Leu is 5667fold lower than wild-type value
E77D
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kcat/Km for beta-Asp-Leu is 137837fold lower than wild-type value
E77Q
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kcat/Km for beta-Asp-Leu is 14571fold lower than wild-type value
R169K
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kcat/Km for beta-Asp-Leu is 378fold lower than wild-type value
R169M
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kcat/Km for beta-Asp-Leu is 1672131fold lower than wild-type value
R233K
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kcat/Km for beta-Asp-Leu is 192fold lower than wild-type value
R233M
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kcat/Km for beta-Asp-Leu is 170fold lower than wild-type value
S289A
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kcat/Km for beta-Asp-Leu is 30000fold lower than wild-type value
T179A
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does not undergo autoprocessing
Y137A
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kcat/Km for beta-Asp-Leu is 927fold lower than wild-type value
Y137F
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kcat/Km for beta-Asp-Leu is 850fold lower than wild-type value
T168A
mutant does not show enzymatic activity
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
medicine
H0VQC8;
isoform ASRGL1 is not the asparaginase responsible for the antitumor effects elicited by treatment with guinea pig serum
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