Information on EC 3.4.11.B4 - Pyrococcus horikoshii deblocking aminopeptidase

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The expected taxonomic range for this enzyme is: Pyrococcus horikoshii

EC NUMBER
COMMENTARY hide
3.4.11.B4
preliminary BRENDA-supplied EC number
RECOMMENDED NAME
GeneOntology No.
Pyrococcus horikoshii deblocking aminopeptidase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
The enzyme exhibits aminopeptidase activity and with lower efficiency deblocking activity (i.e. hydrolysis of acetylated amino acids from the N-terminus of peptides). The turnover number with Ac-Ala-Ala-Ala is 260-540fold lower compared to the activity with Ala-Ala-Ala. The turnover number with Ac-Ala-Ala is 350fold lower compared to the activity with Ala-Ala. Low hydrolytic activity for peptide substrates longer than 10 residues. The enzyme is activated by Co2+ or Zn2+. Contains 1.32 mol of Ca2+ per mol of monomer.
show the reaction diagram
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
physiological function
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Pyrococcus horikoshii has three similar aminopeptidases with deblocking activity. These enzymes appear to play important roles in hydrolyzing small peptides in Pyrococcus horikoshii cells; Pyrococcus horikoshii has three similar aminopeptidases with deblocking activity. These enzymes appear to play important roles in hydrolyzing small peptides in Pyrococcus horikoshii cells
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
Ala-Ala + H2O
L-alanine + L-alanine
show the reaction diagram
Ala-Ala-Ala + H2O
Ala-Ala + L-alanine
show the reaction diagram
Ala-Ala-Ala-Ala + H2O
Ala-Ala-Ala + L-alanine
show the reaction diagram
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-
-
-
?
Ala-Ala-Ala-Ala-Ala + H2O
Ala-Ala-Ala-Ala + L-alanine
show the reaction diagram
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-
-
-
?
N-acetyl-Ala-Ala-Ala + H2O
Ala-Ala + N-acetyl-L-alanine
show the reaction diagram
N-acetyl-L-Ala-Ala + H2O
N-acetyl-L-alanine + L-alanine
show the reaction diagram
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the turnover number with N-acetyl-Ala-Ala is 350fold lower compared to the activity with Ala-Ala
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-
?
N-acetyl-L-alanyl 4-nitroanilide + H2O
N-acetyl-L-alanine + 4-nitroaniline
show the reaction diagram
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-
-
-
?
N-acetyl-L-leucyl 4-nitroanilide + H2O
N-acetyl-L-leucine + 4-nitroaniline
show the reaction diagram
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-
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?
additional information
?
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Zn2+
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activates; activates
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
39.9
Ala-Ala
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pH 7.5, 85C
3.4 - 18.2
Ala-Ala-Ala
13.2
Ala-Ala-Ala-Ala
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pH 7.5, 85C
10.8
Ala-Ala-Ala-Ala-Ala
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pH 7.5, 85C
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.18
Ala-Ala
Pyrococcus horikoshii
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pH 7.5, 85C
0.091 - 9.01
Ala-Ala-Ala
0.022
N-acetyl-L-alanyl 4-nitroanilide
Pyrococcus horikoshii
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pH 7.5, 85C
0.116
N-acetyl-L-leucyl 4-nitroanilide
Pyrococcus horikoshii
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pH 7.5, 85C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.3 - 8.5
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pH 6.3: about 20% of maximal activity, pH 8.5: about 55% of maximal activity; pH 6.3: about 20% of maximal activity, pH 8.5: about 70% of maximal activity
7 - 8.3
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pH 6.5: about 20% of maximal activity, pH 7.0: about 90% of maximal activity, pH 8.3: about 50% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
80 - 110
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80C: about 50% of maximal activity, 110C: about 70% of maximal activity
85 - 110
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85C: about 40% of maximal activity, 110C: about 90% of maximal activity
85 - 98
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85C: about 35% of maximal activity, 98C: about 85% of maximal activity
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
41000
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8 * 41000, SDS-PAGE
42000
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8 * 42000, SDS-PAGE
330000
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gel filtration; gel filtration
440000
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gel filtration, HPLC
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
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x * 36900, wild-type enzyme, mutant enzymes D173N, E205Q and E206Q, SDS-PAGE
octamer
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8 * 41000, SDS-PAGE; 8 * 42000, SDS-PAGE
oligomer
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x * 36900, SDS-PAGE
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
sitting drop method, crystallized in the native and selenomethionine-substituted state, resolution of 3 A; sitting drop method, crystallized in the native and selenomethionine-substituted state, resolution of 3 A; sitting drop method, crystallized in the native and selenomethionine-substituted state, resolution of 3 A
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
90
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2 h, about 20% loss of activity
95
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about 15% loss of activity after 1 h, about 70% loss of activity after 2 h
98
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2 h, over 70% of the activbity remains; 2 h, over 70% of the activity remains
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
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expression in Escherichia coli; expression in Escherichia coli
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overexpressed in Escherichia coli; overexpressed in Escherichia coli; overexpressed in Escherichia coli
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D173N
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Km-value for Ala-Ala-Ala is not significantly different from the KM-value for the wild-type enzyme, kcat is significantly reduced
E205Q
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Km-value for Ala-Ala-Ala is not significantly different from the KM-value for the wild-type enzyme, kcat is significantly reduced
E206Q
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Km-value for Ala-Ala-Ala is not significantly different from the KM-value for the wild-type enzyme, kcat is significantly reduced