Information on EC 3.1.30.1 - Aspergillus nuclease S1

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
3.1.30.1
-
RECOMMENDED NAME
GeneOntology No.
Aspergillus nuclease S1
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
Endonucleolytic cleavage to 5'-phosphomononucleotide and 5'-phosphooligonucleotide end-products
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of phosphoric diester
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hydrolysis of phosphoric ester
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-
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CAS REGISTRY NUMBER
COMMENTARY hide
37288-25-8
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
celery
SwissProt
Manually annotated by BRENDA team
gene ENDO3
UniProt
Manually annotated by BRENDA team
Marburg strain
-
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Manually annotated by BRENDA team
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-
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Manually annotated by BRENDA team
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SwissProt
Manually annotated by BRENDA team
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-
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Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
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-
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Manually annotated by BRENDA team
isolated from colonial ascidium collected near Shikotan Island, Sea of Okhotsk, at a depth of 123 m
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
mung bean
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
the enzyme belongs to the plant S1-like nucleases class of enzymes. Different members of this family are characterized by a surprisingly large variety of catalytic properties, nucleolytic activities of all Arabidopsis thaliana S1-like paralogues, overview. In addition to Zn2+-dependent enzymes, this family also comprises nucleases activated by Ca2+ and Mn2+, which implies that the apparently well-known S1 nuclease active site in plant nucleases is able to cooperate with different activatory ions. Particular members of this class differ in their optimum pH value and substrate specificity. Plant representatives of this family evolve toward an increase in catalytic diversity. Phylogenetic analysis, overview. ENDO3 is the only nuclease, of all members of this family, that demonstrates activity characteristic of fungal S1-type nucleases and mung bean plant nuclease, i.e. it digests ssDNA under acidic pH and in the presence of Zn2+
physiological function
additional information
-
mode of nuclease action and molecular modeling, overview
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
2'-AMP + H2O
adenosine + phosphate
show the reaction diagram
3'-AMP + H2O
adenosine + phosphate
show the reaction diagram
3'-CMP + H2O
cytosine + phosphate
show the reaction diagram
3'-dAMP + H2O
deoxyadenosine + phosphate
show the reaction diagram
3'-dCMP + H2O
deoxycytosine + phosphate
show the reaction diagram
3'-dGMP + H2O
deoxyguanosine + phosphate
show the reaction diagram
3'-dTMP + H2O
deoxythymidine + phosphate
show the reaction diagram
3'-GMP + H2O
guanosine + phosphate
show the reaction diagram
3'-UMP + H2O
uridine + phosphate
show the reaction diagram
adenosine 3'-diphosphate 5'-phosphate
5'-AMP + diphosphate
show the reaction diagram
-
-
-
?
adenosine 3'5'-biphosphate + H2O
5'-AMP + phosphate
show the reaction diagram
ApA + H2O
adenosine + AMP
show the reaction diagram
-
-
-
?
bis(p-nitrophenyl)phosphate + H2O
p-nitrophenol + phosphate
show the reaction diagram
-
almost no hydrolysis
-
?
deoxythymidine 3',5'-bisphosphate
5'-dTMP + phosphate
show the reaction diagram
-
-
-
?
DNA + H2O
5'-phosphomononucleotides + 5'-phosphooligonucleotides
show the reaction diagram
DNA + H2O
?
show the reaction diagram
-
-
-
-
?
DNA + H2O
nicked DNA
show the reaction diagram
dsDNA + H2O
5'-phosphomononucleotides + 5'-phosphooligonucleotides
show the reaction diagram
duplex deoxyadenosine-deoxythymine homopolymer + H2O
5'-dAMP + 5'-TMP + 5'-phosphooligonucleotides
show the reaction diagram
-
-
-
?
heteroduplex DNA + H2O
nicked DNA
show the reaction diagram
platinum-DNA complexes + H2O
5'-phosphomononucleotides + platinum
show the reaction diagram
-
native, denaturated and renaturated platinum-DNA complexes, use of several platinum compounds like diamminedichloroplatinum(II)
-
?
poly(U) + H2O
?
show the reaction diagram
-
preferred substrate
-
-
?
polyA + H2O
fragments of polyA
show the reaction diagram
-
-
-
-
?
polyadenylic acid + H2O
5'-AMP + 5'-phosphooligonucleotides
show the reaction diagram
polycytosylic acid + H2O
5'-CMP + 5'-phosphooligonucleotides
show the reaction diagram
polydeoxyadenylic acid + H2O
5'-dAMP + 5'-phosphooligonucleotides
show the reaction diagram
polydeoxycytosylic acid + H2O
5'-dCMP + 5'-phosphooligonucleotides
show the reaction diagram
-
only little degradation
-
?
polydeoxyguanylic acid + H2O
5'-dGMP + 5'-phosphooligonucleotides
show the reaction diagram
polydeoxythymidylic acid + H2O
5'-dTMP + 5'-phosphooligonucleotides
show the reaction diagram
polyguanylic acid + H2O
5'-GMP + 5'-phosphooligonucleotides
show the reaction diagram
polyinosinic acid + H2O
5'-IMP + 5'-phosphooligonucleotides
show the reaction diagram
-
-
-
?
polyU + H2O
fragments of polyU
show the reaction diagram
-
-
-
-
?
polyuridylic acid + H2O
5'-UMP + 5'-phosphooligonucleotides
show the reaction diagram
RNA + H2O
5'-mononucleotides
show the reaction diagram
-
AMP-preferential endoexonuclease
-
-
?
RNA + H2O
5'-phosphomononucleotides + 5'-phosphooligonucleotides
show the reaction diagram
RNA + H2O
?
show the reaction diagram
-
-
-
-
?
RNA + H2O
fragments of RNA
show the reaction diagram
ssDNA + H2O
5'-phosphomononucleotides + 5'-phosphooligonucleotides
show the reaction diagram
ssDNA + H2O
fragments of ssDNA
show the reaction diagram
tRNA + H2O
5'-phosphomononucleotides + 5'-phosphooligonucleotides
show the reaction diagram
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
DNA + H2O
5'-phosphomononucleotides + 5'-phosphooligonucleotides
show the reaction diagram
DNA + H2O
nicked DNA
show the reaction diagram
heteroduplex DNA + H2O
nicked DNA
show the reaction diagram
poly(U) + H2O
?
show the reaction diagram
-
preferred substrate
-
-
?
RNA + H2O
5'-phosphomononucleotides + 5'-phosphooligonucleotides
show the reaction diagram
RNA + H2O
fragments of RNA
show the reaction diagram
ssDNA + H2O
fragments of ssDNA
show the reaction diagram
additional information
?
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Cd2+
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only marginal stimulating effects
Hg2+
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maximal activity at 0.01-1 mM at pH 3.5-5.0
KCl
-
required
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1,10-phenanthroline
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0.24 mM, complete inhibition of plasmid nicking activity
1-ethyl-3-(3-dimethylaminopropyl)carbodiimide
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inactivation
2-mercaptoethanol
5'-ATP
5'-dAMP
5'-dATP
8-hydroxychinoline 5-sulfonate
amino group modification
aurintricarboxylic acid
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complete inhibition at 0.005 mM
beta-mercaptoethanol
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carboxylate group modification
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at pH 4.6, 50-85% loss of initial activity towards ssDNA, RNA and 3'-AMP after modification of carboxylate groups with 5-15 mM 1-ethyl-3-(dimethylaminopropyl)-carbodiimide; at pH 7.8, 55-95% loss of initial activity towards ssDNA, RNA and 3'-AMP after modification of carboxylate groups with 2-15 mM Woodward's reagent K
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citraconylation
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reversible block of amino groups, lysine residues, by citraconic anhydride
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citrate
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at 0.2 M 50% of initial activity with ssDNA
Co2+
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almost complete inhibition of 3'-AMP hydrolysis by 1 mM CoCl2
CsCl
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optimal activity 0.5-1.8 M, 27% of maximal activity at 7 M
diethyldicarbonate
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inactivation
diphosphate
dithiothreitol
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EGTA
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1 mM, slight inhibition
glutathione
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with 4 mM of the reduced form 50% of activity with denaturated DNA
iodoacetamide
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Ionic strength
MgCl2
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50 mM, 25% inhibition
Mn2+
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6.5% of maximal activity with 0.1 mM denaturated DNA, 8.4% with 0.5 mM polydeoxythymidylic acid as substrate, competitive inhibition
N-bromosuccinimide
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DNA and RNA protects from inactivation
N-ethylmaleimide
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with 5 mM 67% reduction of hydrolysis of polydeoxythymidylic acid
phosphate
Polyvinyl sulfate
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potassium citrate
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complete inhibition at 10 mM
reductive methylation
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modification of lysine residues into N,N'-dimethyl lysine
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sodium bisphosphate
sodium dodecylsulfate
Urea
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40% of maximal activity with single-stranded DNA with 6.6 M
additional information
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urea does not influence the enzyme at any concentrations
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
dimethyl sulfoxide
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glutathione
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oxidized form, about 2fold stimulation of activity at all pH
glycerol
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2fold stimulation of activity by 10-20% in 50 mM Tris-HCl-buffer, pH 7.5
NaCl
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optimal activity at 100 mM
PEG 4000
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Triton X-100
Trypsin
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almost 2.5fold stimulation of activity, probably due to a conversion into a smaller enzyme form
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
208
2'-AMP
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0.04 - 37.6
3'-AMP
71.2
3'-CMP
-
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92
3'-dAMP
-
-
470
3'-dCMP
-
-
223
3'-dGMP
-
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608
3'-dTMP
-
-
47.1
3'-GMP
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-
79.3
3'-UMP
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-
3.4
adenosine 3',5'-bisphosphate
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-
0.02 - 0.7
denaturated DNA
69.9
deoxythymidine 3',5'-bisphosphate
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-
0.0142
DNA
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-
0.02
polydeoxythymidylic acid
-
-
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0.144
RNA
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Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.5
5'-ATP
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-
0.085
5'-dAMP
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0.001
5'-dATP
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.01
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purified enzyme, polydeoxythymidylic acid as substrate
0.63
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with 3'-dAMP as substrate
1.06
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with 2'-AMP as substrate
2.39
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with 3'-dGMP as substrate
3.76
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with 3'dCMP as substrate
4.83
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with 3'-dTMP as substrate
7.7
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mitochondrial activity with ssDNA as substrate
8.3
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purified enzyme, DNA as substrate
12.7
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purified enzyme, heat-denaturated DNA as substrate
17.2
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with deoxythymidine 3',5'-bisphosphate as substrate
241
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with 3'-GMP as substrate
242
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with adenosine 3',5'-bisphosphate as substrate
248
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with 3'-AMP as substrate
255
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with 3'-UMP as substrate
266
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with 3'-CMP as substrate
810
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with heat-denaturated DNA at pH 8.0 in the presence of 10 mM Mg2+
41250
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purified enzyme, pH 3.7, 75C
additional information
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0.6-0.8 U/mg, 1 unit is defined as the amount of enzmyme that is needed to convert 0.001 mg of supercoiled pUC(AT) to either the linear or nicked form in a 0.02 ml reaction at 37C in 30 min
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3.8 - 4.3
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pH 3.8 for the immobilized, pH 4.3 for the soluble enzyme
4.4 - 4.6
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-
4.5 - 5
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with denaturated DNA in the presence of 1 mM Zn2+
6 - 7
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with 3'-AMP and adenosine 3',5'-biphosphate as substrate
6.5
-
cleavage of single base pair mismatches
7.2 - 7.6
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with DNA and RNA as substrate
7.5 - 8.5
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0.1-0.2 M Tris-HCl buffer
additional information
optimal activity at acidic pH
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
2.5 - 8
-
activity range
3 - 8
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almost 50% of activity at pH 3.0 and 5.5
3 - 8.5
-
with mononucleotide substrates
3.1 - 6.4
-
almost 50% of activity at pH 3.3 and 4.6
4 - 6
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with synthetic homopolynucleotides as substrate
4.5 - 10.5
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12.3% of maximal activity at pH 4.5, 43.3% at pH 10.5
5.8 - 9.3
6.5 - 8.5
-
-
6.5 - 8
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6.5 - 7.5
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50% of optimal activity towards ssDNA and dsDNA
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
22
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assay at room temperature
50
-
higher activity at 50C compared to 37C
65
-
immobilized enzyme
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20 - 50
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with tRNA as substrate
20 - 80
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30 - 60
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optimal temperature for denaturated DNA 60C, 13% of maximal activity at 30C
37 - 50
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assay at
47 - 62
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maximal hydrolysis rate of denaturated DNA and RNA
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
9.5
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deduced from nucleotide sequence
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
expression of ENDO3 is highly specific to flower development. It appears first in the shoot apex of an emerging inflorescence stem, remains high in young flowers and decreases gradually with the development of siliques. ENDO3 is preferentially expressed in the pistil and less abundantly in the stamen. The comparison of ENDO3 transcript levels in the whole pistil and separate ovules suggests that ENDO3 activity is mainly related to the ovule
Manually annotated by BRENDA team
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pharyngal carcinoma cell line
Manually annotated by BRENDA team
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microplasmodium
Manually annotated by BRENDA team
-
germinating
Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20000
-
gel filtration
32000 - 38000
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35000: enzyme from nonsenescent flag leaves, 38000: from senescent leaves
34000 - 35000
35000
-
gel filtration
36000 - 38000
-
SDS-PAGE
36000
-
gel electrophoresis
37400
-
SDS-PAGE
38000
-
SDS-PAGE
42000 - 50000
-
-
44000
-
gel filtration
45000
-
SDS-PAGE
47000
-
gel filtration
60000
-
SDS-PAGE
76000
-
-
90000
-
single inactive precursor polypeptide, generation of four nucleases by proteolysis: 75000: Mg2+ dependent exonuclease, 65000: endo-exonuclease, 55000: single-strand-specific endonuclease, 65000: Dnase A, a Ca2+ dependent endonuclease
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
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1 * 35000, SDS-PAGE
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycoprotein
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
crystal structure at 2.1 A resolution
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3.6 - 5.2
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comparatively higher stability of the immobilized than the soluble enzyme
134974
4.8 - 5.5
-
Zn2+ essential for stabilizing activity
134971
5 - 8
-
-
134971
5.5 - 9
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in 30 mM buffer with 200 mM NaCl, more stable above pH 7.0 than below
134988
7 - 8
-
stable without additions
134987
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 70
50
-
40% loss of activity with polyuridylic acid or polydeoxythymidylic acid as substrate after heating of the enzyme fraction for 5 min
55 - 65
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no inactivation by heating at 55C, partial inactivation at 65C, importance of high content of hydrophobic amino acids and zinc atoms for thermostability postulated
60 - 80
-
at 70C complete loss of activity of the soluble enzyme, 40% of maximal activity of the immobilized enzyme at this temperature, 20% at 80C
60 - 67
-
stable below 60C, 50% inactivation after heating at 67C for 15 min, pH 6.0
60
-
after 90 min incubation time 90% of activity towards 3'-AMP and 60% of activity towards DNA lost
60 - 70
-
stable to heating at pH 5.0 in the presence of both Zn2+ and sulfhydryl compounds
65
-
no inactivation of ribonuclease and deoxyribonuclease activity by heating at this temperature in the presence of substrate
74
-
in 0.02 M potassium phosphate buffer, pH 6.5, 37% of activity towards dsDNA after heating of 2 min, no activity after heating of 10 min; in 0.02 M potassium phosphate buffer, pH 6.5, 44% of activity towards ssDNA after heating of 2 min, 4% after heating of 10 min
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
no loss of activity in the presence of 100-250 mM glyoxal
-
stable to high concentrations of urea, susceptible to low concentrations of SDS and guanidine-HCl
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ORGANIC SOLVENT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimethyl sulfoxide
-
no loss of activity in 50% dimethylsulfoxide
dimethylformamide
-
no loss of activity in 30% dimethylformamide
formaldehyde
-
no loss of activity in 2% formaldehyde
formamide
-
no loss of activity in 60% formamide
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, 50% glycerol v/v, 6 months, no loss of activity of the purified enzyme
-
-20C, purified enzyme after dialysis against deionized water, no loss of activity
-
4C, 0.1 M buffer pH 3.5-4.0, mercaptoethanol, 22 h, 16.9% of initial activity at pH 3.5, 59% at pH 4.0, by preincubation with Zn2+ 48% of initial activity at pH 3.5, 86% at pH 4.0
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4C, 30 mM sodium acetate buffer pH 4.6, containing 1 mM ZnSO4, 50 mM NaCl and 5% glycerol, more than 45 days, no apparent loss of activity
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4C, crude and purified form, no loss of activity after several years
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
affinity purification
extracellular enzyme 165fold by chromatography with modified chitosan
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from mitochondria
-
from nucleoplasm
-
heparin-Sepharose, partially purified
-
recombinant carboxy-terminal hexahistidine tag fusion protein
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recombinant endonuclease I
-
recombinant T7 endonuclease
-
recombinant T7 endonuclease I
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recycling isoelectric focusing
-