Information on EC 2.3.1.136 - polysialic-acid O-acetyltransferase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
2.3.1.136
-
RECOMMENDED NAME
GeneOntology No.
polysialic-acid O-acetyltransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
acetyl-CoA + an alpha-2,8-linked polymer of sialic acid = CoA + polysialic acid acetylated at O-7 or O-9
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Acyl group transfer
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
acetyl-CoA:polysialic-acid O-acetyltransferase
Acts only on substrates containing more than 14 sialosyl residues. Catalyses the modification of capsular polysaccharides in some strains of Escherichia coli.
CAS REGISTRY NUMBER
COMMENTARY hide
116412-21-6
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain K1
-
-
Manually annotated by BRENDA team
strain RS174, serotype K1, gene neuO
Q58WP5
SwissProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
-
the homotrimeric enzyme belongs to the left-handed beta-helix family of acyltransferases and is characterized by an unusual funnel-shaped outline
physiological function
-
K1 capsule O-acetylation undergoes highfrequency phase-variation involving also the polysialic acid specific O-acetyltransferase
additional information
-
the enzymatic activity linearly increases with the length of the N-terminal poly-psi-domain which is composed of a variable number of tandem copies of an RLKTQDS heptad, the poly-psi-domain is not resolved in the crystal structure
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
acetyl-CoA + (-8-Neu5Ac-alpha2)2
CoA + ?
show the reaction diagram
-
-
-
-
?
acetyl-CoA + (-8-Neu5Ac-alpha2)3
CoA + ?
show the reaction diagram
-
-
-
-
?
acetyl-CoA + (-8-Neu5Ac-alpha2)4
CoA + ?
show the reaction diagram
-
-
-
-
?
acetyl-CoA + (-8-Neu5Ac-alpha2)5
CoA + ?
show the reaction diagram
-
-
-
-
?
acetyl-CoA + ([Glc-(alpha1-4)-Sia])
?
show the reaction diagram
relative activity: 19.2%
-
-
?
acetyl-CoA + ([Glc-(alpha1-4)-Sia]n)
?
show the reaction diagram
relative activity: 100%
-
-
?
acetyl-CoA + an alpha-2,8-linked polymer of sialic acid
CoA + silaic acid acetylated at O-7 or O-9
show the reaction diagram
acetyl-CoA + CMP-Neu5Ac
CoA + ?
show the reaction diagram
-
-
-
-
?
acetyl-CoA + colominic acid
?
show the reaction diagram
relative activity: 2.1%
-
-
?
acetyl-CoA + colominic acid
CoA + O-(acetyl)-colominic acid
show the reaction diagram
acetyl-CoA + Neu5Ac
CoA + O-(acetyl)-Neu5Ac
show the reaction diagram
-
-
-
-
?
acetyl-CoA + sialic acid polymer of length 12-14
CoA + O-acetyl sialic acid polymer
show the reaction diagram
-
-
-
-
?
Neisseria meningitidis serogroup C capsular polysaccharide + acetyl-CoA
?
show the reaction diagram
-
relative activity: 100%
-
-
?
Neisseria meningitidis serogroup C capsular polysaccharide + propionyl-CoA
?
show the reaction diagram
-
relative activity: 21.4%
-
-
?
propionyl-CoA + colominic acid
CoA + O-(propionyl)-colominic acid
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
acetyl-CoA + an alpha-2,8-linked polymer of sialic acid
CoA + silaic acid acetylated at O-7 or O-9
show the reaction diagram
acetyl-CoA + sialic acid polymer of length 12-14
CoA + O-acetyl sialic acid polymer
show the reaction diagram
-
-
-
-
?
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
acetyl-CoA
-
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
-
no requirement of a divalent cation
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.011
([Glc-(alpha1-4)-Sia]n)
-
0.21 - 0.4
acetyl-CoA
1 - 1.17
colominic acid
0.22
propionyl-CoA
-
mutant NeuO + 36 hepta-nucleotide repeats
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2.1
([Glc-(alpha1-4)-Sia]n)
Neisseria meningitidis
O33391
-
0.51 - 1.3
acetyl-CoA
1.85 - 3.84
colominic acid
0.09
propionyl-CoA
Escherichia coli
-
mutant NeuO + 36 hepta-nucleotide repeats
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
190
([-6)-Glc-(alpha1-4)-Sia-(alpha2-]n) oder ([-6)-Glc-(alpha1-4)-Sia-(alpha2-]n)
Neisseria meningitidis
O33391
-
190107
6.3
acetyl-CoA
Neisseria meningitidis
O33391
-
29
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
-
enzymatic activity increases linearly with increasing numbers of repeats
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7 - 7.5
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-
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
additional information
-
eventually membrane-bound or membrane-associated protein
-
Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
26800
-
SDS-PAGE
54300
-
SDS-PAGE
105000
-
gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hexamer
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gel filtration
homodimer
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SDS-PAGE, the first 34 amino acids form an efficient oligomerization domain, 2 * 54000 Da
homotrimer
additional information
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified engineered apo-NeuO, a protein variant with four N-terminal RLKTQDS heptads, sitting drop vapour diffusion, 6.3 mg/ml protein in 10 mM Tris-HCl pH 7.5 and 150 mM NaCl, mixed with precipitant solution containing 0.1 M Tris, pH 7.2, 0.45 M glycine, 0.2 M NH4NO3, 7% PEG 4000 w/v, 1 week, 20°C, X-ray diffraction structure determination and analysis at 1.7 A resolution, molecular replacement
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OatWY is crystalised with bound substrate acetyl-CoA and its analogs (CoA, S-(2-oxopropyl)-CoA) using the hanging drop vapor diffusion technique. The structure of OatWY reveals an intimate homotrimer of left-handed beta-helix motifs that frame a deep active site cleft selective for the polysialic acid-bearing substrate. A significant movement of Tyr-171 blocks the active site of the enzyme in its native form
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
60
-
stable for 1 h
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
endogenous polysialic acid stabilizes by a close non-covalent association with the enzyme
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Triton X-100, 0.05%, stabilizes
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
4°C, at least 6 months
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
recombinant engineered C-terminally His6-tagged NeuO mutant from Escherichia coli strain BL21 gold (DE3) by nickel affinity chromatography, gel filtration, and ultrafiltration
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using a HisTrap HP column and a a Superdex 200 HR 10/30 column. FInal yield: 24 mg of enzyme from 500 ml of bacterial culture
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using affnitiy chromatography and ion-exchange chromatography
using Ni2+-affinity chromatography
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli as a C-terminal hexahistidine tagged fusion protein
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expressed in Escherichia coli as a C-terminal hexahistidine-tagged fusion protein
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expressed in Escherichia coli as a His-tagged fusion protein
expression of the engineered C-terminally His6-tagged NeuO mutant in Escherichia coli strain BL21 gold (DE3)
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gene neuO, allelic variants, DNA and amino acid sequence determination and analysis, the enzyme is genetically linked to the endo-neuraminidase tail protein gene of a chromosomal accretion element CUS-3, genetic structure, role of heptanucleotide repeat regions, overview
Q58WP5
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
H119A
-
complete loss of enzymatic activity without affecting the oligomerization state
NeuO + 12 hepta-nucleotide repeats
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a set of NeuO alleles with N-terminal 0, 12, 24, and 36 hepta-nucleotide repeats are generated: protein yield: 3.7 mg/L, Km (mM): 0.22 (acetyl-CoA), 1.1 (colomnic acid), kcat (1/sec): 0.56 (acetyl-CoA), 1.85 (colomnic acid)
NeuO + 24 hepta-nucleotide repeats
-
a set of NeuO alleles with N-terminal 0, 12, 24, and 36 hepta-nucleotide repeats are generated: protein yield: 5.0 mg/L, Km (mM): 0.27 (acetyl-CoA), 1.0 (colomnic acid), kcat (1/sec): 1.07 (acetyl-CoA), 2.57 (colomnic acid)
NeuO + 36 hepta-nucleotide repeats
-
a set of NeuO alleles with N-terminal 0, 12, 24, and 36 hepta-nucleotide repeats are generated: protein yield: 5.0 mg/L, Km (mM): 0.22 (acetyl-CoA), 0.22 (propionyl-CoA) 1.6 (colomnic acid), kcat (1/sec): 1.16 (acetyl-CoA), 0.09 (propionyl-CoA), 3.84 (colomnic acid)
W143A
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complete loss of enzymatic activity without affecting the oligomerization state
D359A
-
mutant shows enzymatic activity comparable to wild-type
D371A
-
mutant shows enzymatic activity comparable to wild-type
D376A
-
mutant shows no enzymatic activity
DELTA1-103
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mutant lacking the first 103 amino acids: after purification mutant is found as a monomer, indicating that dimerization of OatC depends on the first 34 amino acids. Mutant shows no activity
DELTA1-34
-
mutant lacking the first 34 amino acids: after purification mutant is found as a monomer, indicating that dimerization of OatC depends on the first 34 amino acids. Mutant retains 25% activity
E336A
-
mutant shows enzymatic activity comparable to wild-type
E345A
-
mutant shows enzymatic activity comparable to wild-type
E379A
-
mutant shows enzymatic activity comparable to wild-type
H111A
-
mutant shows enzymatic activity comparable to wild-type
H121A
mutant shows dramatically reduced activity
H183A
-
mutant shows enzymatic activity comparable to wild-type
H267A
-
mutant shows enzymatic activity comparable to wild-type
H31A
-
mutant shows enzymatic activity comparable to wild-type
H399A
-
mutant shows no enzymatic activity
H456A
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mutant shows reduced enzymatic activity, 70% of wild-type level
S285A
-
mutant retains 80% of wild-type activity
S286A
-
mutant shows no enzymatic activity
S286C
-
mutant shows no enzymatic activity
W145A
mutant shows dramatically reduced activity
Y171A
mutant shows dramatically reduced activity
additional information