Information on EC 3.4.24.33 - peptidyl-Asp metalloendopeptidase

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The expected taxonomic range for this enzyme is: Gammaproteobacteria

EC NUMBER
COMMENTARY
3.4.24.33
-
RECOMMENDED NAME
GeneOntology No.
peptidyl-Asp metalloendopeptidase
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REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
Cleavage of Xaa-/-Asp, Xaa-/-Glu and Xaa-/-cysteic acid bonds
show the reaction diagram
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-
-
-
REACTION TYPE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
hydrolysis of peptide bond
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-
-
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SYNONYMS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
Asp-N endoproteinase
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AspN
Pseudomonas fragi mutant
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-
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AspN endoproteinase
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endoprotease Asp-N
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Endoproteinase Asp-N
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Endoproteinase Asp-N
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Endoproteinase Asp-N
Pseudomonas fragi mutant
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endoproteinase AspN
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Peptidyl-Asp metalloproteinase
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-
-
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Proteinase, peptidyl-Asp metallo-
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-
-
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X-Asp metalloendopeptidase
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CAS REGISTRY NUMBER
COMMENTARY
55576-49-3
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ORGANISM
COMMENTARY
LITERATURE
SEQUENCE CODE
SEQUENCE DB
SOURCE
mutant strain
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-
Manually annotated by BRENDA team
mutant; obtained by growth of Pseudomonas fragi ATCC 4973 on elastin as sole C-source
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Manually annotated by BRENDA team
mutant; strain Me1
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-
Manually annotated by BRENDA team
Pseudomonas fragi Me1
strain Me1
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-
Manually annotated by BRENDA team
Pseudomonas fragi mutant
mutant strain
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-
Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                      
REACTION DIAGRAM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
alpha-Tubulin + H2O
?
show the reaction diagram
-
specific for preaspartate cleavage
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-
-
antithrombin + H2O
?
show the reaction diagram
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the Asp342 localized in helix I is the AspN cleavage site, found in both heating-induced and citrullination-induced polymers of antithrombin
-
-
?
Apolipoprotein A-I + H2O
Peptides
show the reaction diagram
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2 CNBr-fragments, cleavage at 12 Asp-residues and 5 out of 18 Glu-residues, cleaves N-terminal to Glu as well as to Asp and cysteic acid
amino acid sequences
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Azocoll + H2O
?
show the reaction diagram
-
-
-
-
-
casein + H2O
?
show the reaction diagram
-
-
-
-
-
casein + H2O
?
show the reaction diagram
-
resorufin-labeled casein, specific for preaspartate cleavage
-
-
-
Erythrocyte carbonic anhydrase I + H2O
?
show the reaction diagram
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i.e. EC 4.2.1.1, cleavage at 5 Asp- and 1 Glu-residues
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-
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Glucagon + H2O
?
show the reaction diagram
-
-
-
-
?
Glucagon + H2O
?
show the reaction diagram
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specific for preaspartate cleavage
-
-
-
Hemoglobin + H2O
?
show the reaction diagram
-
-
-
-
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Human erythrocyte D-aspartyl-L-isoaspartyl methyltransferase isozyme I + H2O
?
show the reaction diagram
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i.e. EC 2.1.1.77, cleavage sites: N-terminal side of Asp and 5 out of 9 Glu-residues
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Insulin A-chain + H2O
?
show the reaction diagram
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-
-
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Insulin B-chain + H2O
?
show the reaction diagram
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specific for preaspartate cleavage
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Insulin B-chain + H2O
?
show the reaction diagram
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oxidized with performic acid, cleavage sites: Leu6-Cys7, Leu15-Tyr16, Val18-Cys19, Phe24-Phe25
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Myoglobin + H2O
?
show the reaction diagram
Pseudomonas fragi, Pseudomonas fragi mutant
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AspN and 0.5 microg/microl myoglobin at an enzyme to substrate ratio of 1:70 w/w
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-
?
Pancreatic ribonuclease + H2O
Peptides
show the reaction diagram
Pseudomonas fragi, Pseudomonas fragi Me1
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oxidized with performic acid, cleavage sites: -Xaa-Asp- and -Xaa-Cys- (not Cys40, Cys84 or Glu-residues)
i.e. peptide(1-13), peptide(14-25), peptide(26-37), peptide(38-52), peptide(53-57), peptide(53-64), peptide(65-71), peptide(72-82), peptide(83-94), peptide(110-120), peptide(121-124)
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peroxiredoxin II + H2O
?
show the reaction diagram
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-
-
-
?
recombinant histone H1.3 + H2O
C-terminal fragment N.1 of recombinant histone H1.3 + ?
show the reaction diagram
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-
-
-
?
Sperm whale myoglobin + H2O
?
show the reaction diagram
Pseudomonas fragi, Pseudomonas fragi Me1
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in the presence of 2 M urea cleavage of 4 out of 6 Asp-residues
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substrate alpha-parvalbumin + H2O
?
show the reaction diagram
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-
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?
Insulin B-chain + H2O
?
show the reaction diagram
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also cleaves bonds with cysteic acid in P1' derived from cysteine residues by oxidation with performic acid and at N-terminal side of some glutamyl residues
-
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additional information
?
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cleavage specificity
-
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additional information
?
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the wild-type protease has no well-delineated specificity, some preference for N-terminal side of hydrophilic residues, e.g. aminoethylcysteine, Ser, Thr, Gln and Gly
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additional information
?
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endoprotease Asp-N selectively cleaves aspartyl peptides but not the isoaspartyl counterparts
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additional information
?
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high substrate specificity of AspN, that ensures that all of the non-N-terminal peptides having aspartic acid or glutamic acid at their N-termini can be converted. An artificially targeted N-blocked protein is digested with AspN, method overview. The proposed method is applicable to proteins, whether N blocked or N free
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METALS and IONS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
additional information
-
data obtained from inhibition experiments support metalloproteinase character
INHIBITORS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
1,10-phenanthroline
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Zn2+ does not restore after prolonged incubation
1,10-phenanthroline
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cannot be reactivated by Zn2+
acetonitrile
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1% v/v, activation at 5-10% v/v
Alpha-macroglobulin
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inhibition at a molar ratio of inhibitor to protease of about 18 to 1
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additional information
-
not inhibited by PMSF
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ACTIVATING COMPOUND
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
acetonitrile
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slight activation at 5-10% v/v, inhibits at 1%
pH OPTIMUM
pH MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
8
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assay at
TEMPERATURE OPTIMUM
TEMPERATURE OPTIMUM MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
SOURCE
Pseudomonas fragi Me1
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-
-
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
Pseudomonas fragi Me1
-
-
-
-
Manually annotated by BRENDA team
MOLECULAR WEIGHT
MOLECULAR WEIGHT MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
24440
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laser-desorption mass spectrometry
27000
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Pseudomonas fragi, HPLC gel filtration
27000
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SDS-PAGE
additional information
-
-
partial amino acid sequences
SUBUNITS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
monomer
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1 * 27000, Pseudomonas fragi, SDS-PAGE
GENERAL STABILITY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
Stable in up to 2 M urea, 0.01% w/v SDS or 0.1 M guanidine hydrochloride
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ORGANIC SOLVENT
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
acetonitrile
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stable in 5-10% v/v acetonitrile
STORAGE STABILITY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
4°C, lyophilized in the presence of Tris-HCl buffer, pH 7.5, at least 2 years, 4 mg lyophilizate in 100 ml H2O, 2 weeks
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Purification/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
to near homogeneity
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Cloned/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
proteolytic digestion of alpha-parvalbumin using endoproteinase Asp-N for subsequent examination by MALDI-TOF
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ENGINEERING
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
additional information
-
derepressed mutant of Pseudomonas fragi ATCC 4973 produces 40 times higher proteinase levels
APPLICATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
analysis
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analysis of isoaspartic acid by selective proteolysis with Asp-N and electron transfer dissociation mass spectrometry, overview. IsoAsp formation and repair is central to the survival and germination of plant seeds. Also once administered into patients and thus exposed to physiological conditions of pH 7 and 37 °C, protein pharmaceuticals, particularly those with long circulation time, may generate significant amount of isoAsp
molecular biology
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many eukaryotic proteins are blocked at the alpha-amino group of their N-terminal with various modifications, thereby making it difficult to determine their N-terminal sequence by protein sequencer, development of a method for selectively isolating the blocked N-terminal peptide from the peptide mixture generated by endoproteinase AspN digestion of N-blocked protein by removal of all peptides other than the N-terminal one (non-N-terminal peptides) through their carbonyl group introduced by a chemical transamination reaction
additional information
-
AspN is shown to be an alternative protease for in-capillary digestion (during capillary electrophoresis), AspN behaves very similarly to trypsin
additional information
Pseudomonas fragi mutant
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AspN is shown to be an alternative protease for in-capillary digestion (during capillary electrophoresis), AspN behaves very similarly to trypsin
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