Information on EC 3.1.14.1 - yeast ribonuclease

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
3.1.14.1
-
RECOMMENDED NAME
GeneOntology No.
yeast ribonuclease
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
exonucleolytic cleavage to nucleoside 3'-phosphates
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of phosphoric ester
-
-
-
-
phosphoric ester hydrolysis
CAS REGISTRY NUMBER
COMMENTARY hide
9001-99-4
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
Saccharomyces fragilis x Saccharomyces dobzhanski
-
-
-
Manually annotated by BRENDA team
gene pnu1+
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
separable components RNase U4A, U4B, U4C
-
-
Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
DNA + H2O
deoxynucleoside 3'-phosphates
show the reaction diagram
-
pUC18 or pSP1
-
?
polyadenylic acid + H2O
3'-AMP
show the reaction diagram
Saccharomyces fragilis x Saccharomyces dobzhanski
-
-
-
?
polyuridylic acid + H2O
3'-UMP
show the reaction diagram
Saccharomyces fragilis x Saccharomyces dobzhanski
-
-
-
?
ribonucleic acid + H2O
3'-phosphomononucleotides
show the reaction diagram
RNA + H2O
nucleoside 3'-phosphate + ?
show the reaction diagram
-
in vitro transcribed tRNAGlu3 UUC, tRNAGIn UUG and tRNALys UUU are substrates, stable anticodon stem and the anticodon loop are the minimal requirements for cleavage, synthetic minihelix RNA corresponding to the anticodon stem loop of the natural substrate tRNAGlu3 mcm5s2UUC is cleaved at the same position as the natural substrate. In ASLGlu3 UUC, the nucleotides U34U35C36A37C38 are required for optimal gamma-toxin cleavage, whereas a purine at position 32 or a G in position 33 dramatically reduces the cleavage of the anticodon stem loop
-
-
?
RNA + H2O
nucleoside 3'-phosphates
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
ribonucleic acid + H2O
3'-phosphomononucleotides
show the reaction diagram
RNA + H2O
nucleoside 3'-phosphates
show the reaction diagram
-
-
-
?
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
KH2PO4
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10-80% activation at various concentrations
Na2SO4
-
slight activation
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1,4-dithiothreitol
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50% loss of activity at 5 mM, 75% at 10 mM
Ag+
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no activity at 1 mM AgNO3
Ba2+
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85% of maximal activity at 1 mM BaCl2
Ca2+
-
77% of activity at 1 mM CaCl2, 73% at 10 mM
EDTA
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recoverage by either Mg2+, Cl-, or CaCl2
Fe2+
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76% of activity at 1 mM FeCl2, 46% at 10 mM
Fe3+
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41% of maximal activity at 1 mM FeCl3
glutathione
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95% inhibition by 0.01 M
Hg2+
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no activity at 1 mM HgCl2
Mg2+
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91% of activity at 1 mM MgCl2, 77% at 10 mM
N-bromosuccinimide
-
-
Polyvinyl sulfate
Saccharomyces fragilis x Saccharomyces dobzhanski
-
70% of maximal activity with 50 microg/ml, 10% of maximal activity with 100 microg/ml
spermidine
Saccharomyces fragilis x Saccharomyces dobzhanski
-
slight inhibition: 80% of maximal activity with 0.01 M
additional information
-
tryptophan residues and disulfide bonds required for activity
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
phenylmethanesulfonyl fluoride
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0.1 mM: inhibition of intracellular proteolytic proteins during the incubation of disrupted yeast cells for RNA hydrolysis
protease isp6+
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strongly dependent on, gene isp6+ encodes a protease, which is involved in RNA-degrading enzyme activity, an isp6+-deficient mutant is also devoid of RNA-degrading enzyme activity
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Trimethylamine N-oxide
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enhances cleavage of unmodified tRNA and anticodon stem loops by gamma-toxin-GST
additional information
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
11.3
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179fold partial purification
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4 - 4.25
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0.25 M sodium acetate buffer
4.5
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with yeast RNA as substrate
6 - 7
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U4A: 7.0; U4B: 6.5; U4C: 6.0
7
Saccharomyces fragilis x Saccharomyces dobzhanski
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nuclease obtained from stationary or exponentially growing cells
8
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RNA helicase assay at
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3 - 9
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50% of activity near pH 5.0
4 - 7
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the pH does not much influence the RNA degradation activity
4.5 - 9
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Ph 4.5: about 15% of maximum activity, pH 9.0: about 50% of maximum activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50
-
30 min reaction time
50 - 55
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sodium acetate buffer at pH 4.0
55
-
with yeast RNA as substrate
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 60
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the composition of RNA degradation products varies with autolysis temperature, overview
35 - 65
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less than 15% of maximum activity above and below
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
Saccharomyces fragilis x Saccharomyces dobzhanski
-
-
Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20000 - 23000
-
207000 for RNase U4A, 21300 for RNase U4B, 22100 RNase U4C; gel filtration
26000 - 29000
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29500 for RNase U4A, 26900 for RNase U4B, 20000for RNase U4C; equilibrium ultracentrifugation
38000
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x * 38000, SDS-PAGE
56000
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sedimentation equilibrium method
58900
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sedimentation diffusion method
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
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x * 38000, SDS-PAGE
additional information
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4 - 10
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at pH 8 stable up to at least 50C, inactivation at higher temperatures; stable at 37C for 1 h
134106
5 - 10
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at 2C; at 40C: inactivation rate at pH 10 markedly higher than at pH 4.1, 6.3 and 8.6
134105
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
40
-
several h under toluene, pH 7.5
50
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10 min, less than 10% loss of activity
60
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remaining activity after heating at 60C for 90 min at pH 5.4 30%, at pH 7.0 40%, at pH 9.0 50%
70
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2 min, complete inactivation
80
-
almost no activity
additional information
-
NaCl and KCl might protect from heat denaturation at 52C
OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
disulfide bonds required for activity
-
134105
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, 50% glycerol, stable for 12 months
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4C, under toluene, pH 7.5, several weeks, little loss of activity
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
partial purification
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recombinant degradosome from Saccharomyces cerevisiae W303 strain derivatives
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recombinant wild-type enzyme from overexpressing cells
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression of wild-type and mutants in different yeast strains
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gamma-toxin-GST expressed in Escherichia coli BL21(DE3)
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overexpression of wild-type enzyme in Schizosaccharomyces pombe
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Y814S
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point mutation in Dssp1, mutant strain is respiratory incompetent with very unstable mitochondrial genomes
additional information
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construction of a enzyme null-mutant pnu1DELTA, pnu1+ is dispensable for mitochondrial function and not essential for cell growth
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
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knowledge of amino acid sequences are useful for phylogenetic studies of several fungi
nutrition
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possibility for reducing nucleic acid content in yeast protein used in human food