Information on EC 3.1.1.53 - sialate O-acetylesterase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
3.1.1.53
-
RECOMMENDED NAME
GeneOntology No.
sialate O-acetylesterase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
N-acetyl-O-acetylneuraminate + H2O = N-acetylneuraminate + acetate
show the reaction diagram
acts on free and glycosidically bound N-acetyl- or N-glycoloyl-neuraminic acid, acts mainly on the 4-O- and 9-O-acetyl groups. Also acts on some other O-acetyl esters, both cyclic and acyclic compounds, which are not sialic acids
-
-
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
acetylation
hydrolysis of carboxylic ester
-
-
-
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SYSTEMATIC NAME
IUBMB Comments
N-acyl-O-acetylneuraminate O-acetylhydrolase
Acts on free and glycosidically bound N-acetyl- or N-glycoloyl-neuraminic acid; acts mainly on the 4-O- and 9-O-acetyl groups. Also acts on some other O-acetyl esters, both cyclic and acyclic compounds, which are not sialic acids.
CAS REGISTRY NUMBER
COMMENTARY hide
89400-31-7
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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-
-
Manually annotated by BRENDA team
starfish
-
-
Manually annotated by BRENDA team
VIII-271C, VIII-271G, VIII-271E, VIII-271F, isolated from human faeces
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
VIII-210, isolated from human faeces
-
-
Manually annotated by BRENDA team
X-95, isolated from human faeces
-
-
Manually annotated by BRENDA team
VII-240, isolated from human faeces
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
BcoV
-
-
Manually annotated by BRENDA team
sea bass
-
-
Manually annotated by BRENDA team
X-18B, isolated from human faeces
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
ISAV 4, isolate 4, Glesvaer/2/90
-
-
Manually annotated by BRENDA team
INF-C
UniProt
Manually annotated by BRENDA team
influenza C virus Johannesburg/1/66
strain Johannesburg/1/66
-
-
Manually annotated by BRENDA team
MHV-DVIM, a Murine coronavirus, MuCoV, or Mouse hepatitis virus, MHV, species
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-
Manually annotated by BRENDA team
MHV-DVIM, a Murine coronavirus, MuCoV, or Mouse hepatitis virus, MHV, species
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-
Manually annotated by BRENDA team
MHV-S, a Murine coronavirus, MuCoV, or Mouse hepatitis virus, MHV, species
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-
Manually annotated by BRENDA team
no activity in Dictyostelium discoideum
-
-
-
Manually annotated by BRENDA team
no activity in Glycine max
-
-
-
Manually annotated by BRENDA team
no activity in Hirudo medicinalis
medicinal leech
-
-
Manually annotated by BRENDA team
no activity in Nicotiana tabacum
-
-
-
Manually annotated by BRENDA team
no activity in Spodoptera frugiperda
fall armyworm, Sf-9 cells
-
-
Manually annotated by BRENDA team
VI-268, isolated from human faeces
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-
Manually annotated by BRENDA team
IX-70, VIII-239, isolated from human faeces
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-
Manually annotated by BRENDA team
serotype 2
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-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
malfunction
physiological function
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
2-deoxy-2,3-didehydro-N-acetyl-9-O-acetylneuraminate + H2O
?
show the reaction diagram
3'-O-acetylthymidine + H2O
thymidine + acetate
show the reaction diagram
-
8% of the activity compared to N-acetyl-9-O-acetylneuraminate
-
-
?
4-methylumbelliferyl acetate + H2O
4-methylumbelliferone + acetate
show the reaction diagram
4-methylumbelliferyl butyrate + H2O
4-methylumbelliferone + butyrate
show the reaction diagram
4-nitrophenyl acetate + H2O
4-nitrophenol + acetate
show the reaction diagram
4-nitrophenylacetate + H2O
4-nitrophenol + acetate
show the reaction diagram
-
-
-
-
?
4-O-acetyl sialic acid + H2O
sialic acid + acetate
show the reaction diagram
-
-
-
-
?
5-N-acetyl-4,9-di-O-acetylneuraminic acid alpha-methylglycoside + H2O
4,9-di-O-acetylneuraminic acid alpha-methylglycoside + acetate
show the reaction diagram
sialate-9-O-acetylesterase activity
analysis by gas chromatography/mass spectrometry
-
?
5-N-acetyl-7(8),9-di-O-acetylneuraminic acid + H2O
?
show the reaction diagram
-
-
-
?
5-N-acetyl-9-O-acetylneuraminic acid + H2O
5-N-acetylneuraminic acid + acetate
show the reaction diagram
9-O-acetyl sialic acid + H2O
sialic acid + acetate
show the reaction diagram
9-O-acetyl-N-acetylneuraminic acid + H2O
N-acetylneuraminic acid + acetate
show the reaction diagram
-
-
-
-
?
acetyl-CoA + H2O
CoA + acetate
show the reaction diagram
alacepril + H2O
deacetylalacepril + acetate
show the reaction diagram
-
-
-
?
alpha-naphthyl acetate + H2O
1-naphthol + acetate
show the reaction diagram
alpha-naphthyl acetate + H2O
alpha-naphthol + acetate
show the reaction diagram
-
-
-
?
bovine submandibular gland mucin + H2O
?
show the reaction diagram
bovine submaxillary mucin + H2O
?
show the reaction diagram
-
-
-
-
?
CMP-O9-acetyl-N-acetylneuraminate + H2O
CMP-N-acetylneuraminate + acetate
show the reaction diagram
-
SsNeuA is a bifunctional CMP-Neu5Ac synthetase/O-acetylesterase, which strictly de-O-acetylates CMP-O-acetyl-Neu5Ac
-
-
?
cytidine 5'-monophospho-N-acetylneuraminic acid + H2O
?
show the reaction diagram
methyl 6-O-(N-acetyl-9-O-acetyl-alpha-D-neuramin-2-yl)-alpha-D-glucoside + H2O
methyl 6-O-(N-acetyl-alpha-D-neuramin-2-yl)-alpha-D-glucoside + acetate
show the reaction diagram
-
methyl N-acetylneuraminide, 25C, 60 min, pH 7
NMR-based enzyme assay
-
?
methyl 6-S-(N-acetyl-9-O-acetyl-alpha-D-neuramin-2-yl)-6-thio-alpha-D-glucoside + H2O
methyl 6-S-(N-acetyl-alpha-D-neuramin-2-yl)-alpha-D-glucoside + acetate
show the reaction diagram
N-acetyl-4-O-acetylneuraminate + H2O
N-acetylneuraminate + acetate
show the reaction diagram
N-acetyl-4-O-acetylneuraminic acid + H2O
N-acetylneuraminate + acetate
show the reaction diagram
-
pH 8, 30 min, 37C, catalysed only by esterase with pI 5.7
analyses by HPLC
-
?
N-acetyl-9-O-acetyl-neuraminic acid + H2O
N-acetylneuraminic acid + acetate
show the reaction diagram
N-acetyl-9-O-acetyl-neuraminic acid alpha-methylglycoside + H2O
N-acetylneuraminic acid alpha-methylglycoside + acetate
show the reaction diagram
methyl N-acetylneuraminide, 25C, 60 min, pH 7
NMR-based enzyme assay
-
?
N-acetyl-9-O-acetylneuraminate + H2O
N-acetylneuraminate + acetate
show the reaction diagram
N-acetyl-9-O-acetylneuraminate beta-methylglycoside + H2O
N-acetylneuraminate beta-methylglycoside + acetate
show the reaction diagram
-
70% of the activity compared to N-acetyl-9-O-acetylneuraminate
-
-
?
N-acetyl-9-O-acetylneuraminate lactose + H2O
N-acetylneuraminate lactose + acetate
show the reaction diagram
N-acetyl-9-O-acetylneuraminic acid + H2O
?
show the reaction diagram
-
-
-
-
?
N-acetyl-9-O-acetylneuraminic acid + H2O
N-acetylneuraminate + acetate
show the reaction diagram
-
pH 8, 30 min, 37C, catalysed by esterase with pI 4.8 and esterase with pI 5.7
analyses by HPLC
-
?
N-acetyl-O-9-acetylneuraminate + H2O
N-acetylneuraminate + acetate
show the reaction diagram
N-acetyl-O-acetylneuraminate + H2O
?
show the reaction diagram
-
-
-
-
-
N-acetyl-O-acetylneuraminate + H2O
N-acetylneuraminate + acetate
show the reaction diagram
-
destroys receptors on the surface of the target cells
-
-
?
O-9-acetyl-GD3-S-phenyl + H2O
?
show the reaction diagram
-
i.e. 9-O-acetyl-NeuAcalpha-2,8-NeuAcalpha-2,3-Galbeta-1,4-Glcbeta-S-phenyl
-
-
?
O-acetylserine + H2O
serine + acetate
show the reaction diagram
-
less than 5% of the activity compared to N-acetyl-9-O-acetylneuraminate
-
-
?
p-nitrophenyl acetate + H2O
p-nitrophenol + acetate
show the reaction diagram
rat serum glycoprotein + H2O
?
show the reaction diagram
-
90% of the activity compared to N-acetyl-9-O-acetylneuraminate
-
-
?
spironolactone + H2O
?
show the reaction diagram
-
-
-
?
thiophenyl acetate + H2O
thiophenol + acetate
show the reaction diagram
triacetin + H2O
?
show the reaction diagram
-
26% of the activity compared to N-acetyl-9-O-acetylneuraminate
-
-
?
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
5-N-acetyl-9-O-acetylneuraminic acid + H2O
5-N-acetylneuraminic acid + acetate
show the reaction diagram
CMP-O9-acetyl-N-acetylneuraminate + H2O
CMP-N-acetylneuraminate + acetate
show the reaction diagram
-
SsNeuA is a bifunctional CMP-Neu5Ac synthetase/O-acetylesterase, which strictly de-O-acetylates CMP-O-acetyl-Neu5Ac
-
-
?
N-acetyl-O-9-acetylneuraminate + H2O
N-acetylneuraminate + acetate
show the reaction diagram
-
contribution of Ser19 and His301 to catalysis, active site structure, overview
-
-
?
N-acetyl-O-acetylneuraminate + H2O
?
show the reaction diagram
-
-
-
-
-
N-acetyl-O-acetylneuraminate + H2O
N-acetylneuraminate + acetate
show the reaction diagram
-
destroys receptors on the surface of the target cells
-
-
?
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Cu2+
-
1 mM, block of essential SH-groups
Hg2+
-
1 mM, block of essential SH-groups
Mg2+
-
activates
NaCl
-
150 mM, inhibitory at 500 mM when pH below pH 8
Zn2+
-
1 mM, block of essential SH-groups
additional information
-
no divalent cation requirement
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2,3-Butanedione
2-alpha-thiomethylmercuryl 9-acetamido-9-deoxy sialoside
-
competitive inhibition, Ki: 4.2 mM; the inhibitor is used to prepare heavy atom derivatives of the crystals
3,4-dichloroisocoumarin
9-acetamido-N-acetyl-O-acetylneuraminate
Alpha-naphthyl acetate
-
competitive inhibitor
ammonium (allyl 5-acetamido-3,5-dideoxy-4-O-methyl-D-glycero-alpha-D-galacto-2-nonulopyranosidonate)
ammonium (allyl 5-acetamido-9-O-methyl-3,5-dideoxy-D-glycero-alpha-D-galacto-2-nonulopyranosidonate)
bis-p-nitrophenyl phosphate
-
little effect on the activity
CHAPS
-
lowers the activity by 40%
deoxycholate
-
75-80% loss of activity at 2% concentration
diammonium (allyl 5-acetamido-3,5-dideoxy-4-O-(P-methylphosphonyl)-D-glycero-alpha-D-galacto-2-nonulopyranosidonate)
diammonium (allyl 5-acetamido-3,5-dideoxy-9-O-(P-methylphosphonyl)-D-glycero-alpha-D-galacto-2-nonulopyranosidonate)
Diethyl-4-nitrophenylphosphate
diisopropyl fluorophosphate
diisopropyl fluorophosphates
-
complete inhibition at 1 mM
diisopropylfluorophosphate
-
1 mM, 25C, 15 min, total loss of activity
F-
-
little effect on the activity
N-acetyl-9-O-acetylneuraminate
-
substrate inhibition above 2 mM
p-chloromercuribenzoate
-
little effect on the activity
Paraoxon
-
diethyl-4-nitrohenylphosphate (E600), 1 mM at 25C within 15 min leads to total loss of activity
Phenylglyoxal
phenylmethylsulfonyl fluoride
-
complete inhibition at 10 mM
physostigmine
PMSF
-
45% inhibition at 1 mM
additional information
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
CTP
-
activates
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.3
4-O-acetyl-sialic acid
-
low pI esterase form, obtained with free sialic acid
0.13 - 0.17
4-yethylumbelliferyl acetate
1.1 - 1.3
9-O-acetyl-sialic acid
1.28
acetyl-CoA
-
pH 7.5, 37C
22
alacepril
-
pH 7.5, 37C
1.71
Alpha-naphthyl acetate
-
pH 7.5, 37C
0.8 - 18
N-acetyl-9-O-acetylneuraminic acid
0.9 - 24
N-acetyl-9-O-acetylneuraminic acid lactose
0.652
spironolactone
-
pH 7.5, 37C
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
additional information
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
4.2
2-alpha-thiomethylmercuryl 9-acetamido-9-deoxy sialoside
-
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.00012
-
homogenate
0.05
-
specific activity in the commercial enzyme product Novarom G
0.104
-
after preparative isoelectric focussing, pI 4.8
0.12
-
specific activity in the commercial enzyme product Ultrazym
0.17
-
strain VI-268
0.19
-
strain VIII-239
0.31
-
strain IX-70
0.4
-
specific activity in the commercial enzyme product Vinozym
0.71
-
specific activity in the commercial enzyme product AR2000
0.84
-
4-methylumbelliferyl acetate as substrate
1.29
-
substrate O-9-acetyl-GD3-S-phenyl, pH 7.2, 25C
1.8
-
purified recombinant 29 kDa O-acetylesterase fragment, pH 7.2, 37C
2.31
-
substrate N-acetyl-O-9-acetylneuraminate, pH 7.2, 25C
11.6
-
alpha-naphthyl acetate as substrate
12.8
-
4-nitrophenyl acetate as substrate
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 7.5
-
-
7.2
-
assay at
7.5
-
assay at
8
-
HEPES buffer, activity affected at high NaCl concentrations (500 mM)
8 - 8.5
-
substrate 4-nitrophenyl acetate, wild-type SsNeuA and mutant SsNeuA234-410
8.4 - 8.6
-
-
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 8
-
pH 5: 16% of the activity, pH 8: 80% of the activity
6.8 - 8.2
-
HEPES buffer
7
-
60% lower activity compared to optimum
7 - 8.5
-
using 4-nitrophenyl acetate as substrate
7.5 - 9
-
-
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4.8
-
isoelectric focusing, low pI form
5.7
-
isoelectric focusing, high pI form
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
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high levels of activity of both forms of the enzyme, nonglycosylated and glycosylated
Manually annotated by BRENDA team
-
hepatoietic cell line
Manually annotated by BRENDA team
-
almost exclusively the nonglycosylated form of the enzyme
Manually annotated by BRENDA team
-
high levels of activity of both forms, nonglycosylated and glycosylated
Manually annotated by BRENDA team
-
normal and cancer mucosa
Manually annotated by BRENDA team
-
restricted distribution, strongest expression of the lysosomal enzyme
Manually annotated by BRENDA team
-
only the glycosylated form of the enzyme
Manually annotated by BRENDA team
-
high levels of activity of both forms, nonglycosylated and glycosylated
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
-
isoform Lse, secreted from post-Golgi vesicle, limited lysosomal localisation
Manually annotated by BRENDA team
additional information
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37230
-
MALDI-TOF mass spectrometry
40000
-
SDS-PAGE, purified, recombinant protein
54000
-
gel-filtration
60000
-
gel filtration
62800
-
gel filtration
80000
-
SDS PAGE under non-reducing conditions
88000
-
gel fitration
250000
-
nonreducing PAGE, equal MW for both the larger and smaller Lse subunit, expressed in COS-7 cells
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
heterodimer
monomer
additional information
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycoprotein
side-chain modification
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
apo-enzyme (PDB: 3CL4) or mutant S40A in complex with 5-N-acetyl-4,9-di-O-acetylneuraminic acid alpha-methylglycoside (PDB: 3CL5), hanging drop vapour diffusion, 18C, 2 weeks, precipitant: 16% polyethylene glycol 8000 and 20% glycerol or 10% polyethylene glycol 3350, hexagonal bipyramide crystals: space groups: P6(5)22, unit cell parameter: a, b: 88.8-89.3, c: 280.4-282.4, soaking with 7 mM 5-N-acetyl-4,9-di-O-acetylneuraminic acid alpha-methylglycoside, molecular replacement using PDB: 1FLC (for apo-enzyme) or wild-type structure (for S40A mutant) as template, three domains: receptor-binding domain (R), acetylase domain (E, strictly conserved), and membrane-proximal domain (MP), homodimer of 2fold crystallographic symmetry with 2 major contact regions (CR1: bridges R domains, CR2: involves MP), disorder of residues 377-388, E-domain: SGNH-hydrolase fold, active site with catalytic triad (Ser40, His329, Asp326) and oxyanion hole (Asn104, Ser40, Gly75), highly variable surface loop (residues 47-54) is site of antigenic variation, R-domain: different from those of hemagglutinin-esterase fusion (HEF) and agglutinin (HA) (plasticity), binding of ligand in opposite orientation involving residues Leu212, Asn214, Ser213, Tyr184, Phe211, Leu266, Leu267 (hydrophobic pocket), coordination of potassium and water by Asp220, Ser221, Gln222, Ser263, Glu265, Leu267
purified wild-type apo-NanS and SeMet-labeled NanS, mixing of 0.001 ml of protein in the final buffer with 0.001 ml of reservoir solution containing 0.2 M NaCl, 0.1 M Bis-Tris pH 5.5, 30% w/v PEG 3350, in micro batch experiments under oil, X-ray diffraction structure determination and analysis at 1.6-2.2 A resolution
-
molecular modelling using PDB: 1FLC, and alpha-methyl or beta-methyl glycosides of 9-acetamido-9-deoxy-N-acetylneuraminic acid or N-acetyl-9-O-acetyl-2,6-alpha-S-glucosyl-neuraminic acid alpha-methylglycoside as substrates, no interaction between aglycon moiety and the enzyme or its active site, catalytic triad: Ser57, His355, Asp352, catalysis involves conformational change of Ser57 side chain in order to enable 9-O-acetyl-binding and position for nucleophilic attack on 9-O-acetate carbonyl carbon, beta-anomer substrate lacks bi-dendate interaction between C-1 carboxylate group and Asp352 and is no suitable substrate
the inhibitor 2-alpha-thiomethylmercuryl 9-acetamido-9-deoxy sialoside is used to prepare heavy atom derivatives of the crystals
-
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25
-
room temperature, complete inactivation in the presence of 10 mM 2-mercaptoethanol after 8 h
37
-
40% activity is lost in 20 h
96
-
in the presence of bovine serum albumin, loss of activity within 1 min
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
activity becomes unstable on excessive dilution
-
lyophilization and reconstitution results in loss of 30% of activity
-
partially purified esterase (from second ion exchange chromatography) resistant to freezing-thawing cycles, presence of beta-mercaptoethanol during purification preserves enzyme activity
-
single cycle of freeze-thaw results in little loss of activity, repeated freeze-thaw results in gradual loss of activity
-
OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
15 min incubation with 1 mM 4-hydroxymercuribenzoate at 25C leads to total loss of activity
-
698132
contains disulfide bonds
-
701059
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, 20 mM HEPES 150 mM NaCl pH 8, 3 weeks, fully active
-
-20C, after preparative isoelectric focusing, few weeks, total loss of activity, -70C, frozen liver, stable for 1-2 years
-
-80C, in lyophilized form, no loss of activity
-
37C, in the presence of bovine serum albumin, decrease of 50% of the activity within 2 h
-
4C, 20 mM HEPES 150 mM NaCl pH 8, 1 week, fully active
-
4C, concentrated purified enzyme, stable for at least 1 month
-
4C, Tris-HCl, pH7.8, 2 mM mercaptoethanol, 90% loss of activity after 1 day, addition of 15 mM bovine serum albumin prevents loss during 7 days
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
180fold enrichment
-
2600fold enrichment
-
850fold enrichment
-
copurified with a 66000 Da esterase
-
from baculovirus infected insect Sf9 cells as GFP-fusion protein by ultracentrifugation, ion-exchange chromatography on Q-Sepharose FF column and dialysis
from fresh tissue by ultracentrifugation (dialysis of supernatant), followed by gel-filtration on Sephadex G-200 column, ion-exchange chromatography on Sephadex-DEAE-A-50 column, second ion-exchange chromatography on DEAE-Sephacel column, isoelectric focussing on ampholine of pH 3.5-9.5 and a saccharose gradient, and final dialysis, 866% purification compared to homogenate
-
from HEK293S cells by protein A-affinity chromatography followed by on-bead thrombin digestion
nickel-affinity chromatography, elution with 500 mM imidazole, pH 7.5, yield of 300 microg soluble recombinant protein per litre Sf9 cell culture (3 mg/l insoluble)
-
nickelnitrilotriacetic acid-agarose chromatography
-
separated from alpha-L-arabinofuranosidase and alpha-L-rhamnopyranosidase in commercial enzyme preparations
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed as GFP-fused secretion protein in Sf9 cells
-
expressed in COS-7 cells, both the larger and smaller Lse subunit
-
expression in E. coli BL21
-
expression of wild-type and mutant enzymes in Escherichia coli BL21(DE3). SsneuA can restore both CMP-Neu5Ac synthetase and O-acetylesterase activities in complemented enzyme-deficient Escherichia coli strains, but the activities in the complemented strain are different from those in the wild-type Escherichia coli strain. However, the Streptococcus suis O-acetylesterase domain alone is unable to act on intracellular O-acetyl-Neu5Ac in Escherichia coli
-
genotyping, exome sequencing, in HEK293 T cells overexpressed recombinant enzyme can be secreted by the transfected lymphocates, in the contrary to the physiological state of the enzyme
-
genotyping, exome sequencing, in HEK293 T cells overexpressed recombinant enzyme can be secreted by the transfected lymphocytes, in the contrary to the physiological state of the enzyme
-
in pBACgus-6, -3, -1 for baculovirus-mediated expression for 4 h at 20C with C-terminal hexa-His-tag in insect Sf9 cells, deletion of transmembrane region and cytoplasmic tail
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sequence encoding ectodomain (residues 19-388) in plasmid pCD5-BCoVHE-T-Fc for expression with N-terminal CD5 signal peptide and C-terminal thrombin cleavage site followed by human IgG1 Fc domain in N-acetylglucosaminyltransferase-deficient HEK293S cells
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
expression of lysosomal and cytosolic form of SIAE is significantly downregulated both in lymphoblasts of acute lymphoblastic leukemia patients and acute lymphoblastic leukemia-cell line in comparison to peripheral blood mononuclear cells from healthy donors, semi-quantitative RT-PCR using specific primers for both SIAE genes, overview
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
F211A
abrogated ligand recognition and binding
L266A
abrogated ligand recognition and binding
L267A
abrogated ligand recognition and binding
S40A
catalytically inactive, active site residue, retained lectin activity
Y184A
decreased ligand binding affinity
E26A
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site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type NanS
H301N
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site-directed mutagenesis, inactive mutant
S19A
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site-directed mutagenesis, inactive mutant
S300A
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site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type NanS
T294A
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site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type NanS
T294S
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site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type NanS
A385T
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naturally occuring mutation
A394T
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naturally occuring mutation
C443R
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naturally occuring mutation
F404S
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naturally occuring mutation
G419E
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naturally occuring mutation
G514A
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naturally occuring mutation
G64S
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naturally occuring mutation, the enzyme shows unaltered catalytic activity compared to the wild-type enzyme. The mutant enzyme is poorly secreted when overexpressed
H447R
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naturally occuring mutation
H472Q
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naturally occuring mutation
K434T
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naturally occuring mutation
M456I
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naturally occuring mutation
M456T
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naturally occuring mutation
M89V
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naturally occuring mutation, the enzyme shows unaltered catalytic activity compared to the wild-type enzyme. The mutant enzyme is poorly secreted when overexpressed
Q309P
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naturally occuring mutation, the mutant shows highly reduced catalytic activity compared with the wild-type enzyme
Q428L
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naturally occuring mutation
Q462R
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naturally occuring mutation
R387W
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naturally occuring mutation
R393C
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naturally occuring mutation
R393H
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naturally occuring mutation
R479C
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naturally occuring mutation
S127A
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naturally occuring mutation, the enzyme shows below 10% catalytic activity compared to the wild-type enzyme
V459I
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naturally occuring mutation
S57T
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loss of esterase activity
C196F
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naturally occuring mutation, the mutant shows highly reduced catalytic activity compared with the wild-type enzyme
Q335P
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naturally occuring mutation, the mutant shows highly reduced catalytic activity compared with the wild-type enzyme
S258A
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site-directed mutagenesis
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
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detection of 9-O-acetylated sialic acids on sialoglycoconjugates immobilized on microtiter plates, nitrocellulose or separated on thin-layer chromatograms. The assay takes advantage of two different biological properties of influenza C virus, its high affinity-binding to 9-O-acetylated sialic acids and its sialate 9-O-acteylesterase that is used for detection of bound virus
drug development
molecular biology
origin and evolution of viral hemagglutinin-esterases
Show AA Sequence (631 entries)
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