Information on EC 2.4.1.231 - alpha,alpha-trehalose phosphorylase (configuration-retaining)

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The expected taxonomic range for this enzyme is: Eukaryota, Bacteria

EC NUMBER
COMMENTARY hide
2.4.1.231
-
RECOMMENDED NAME
GeneOntology No.
alpha,alpha-trehalose phosphorylase (configuration-retaining)
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
alpha,alpha-trehalose + phosphate = alpha-D-glucose + alpha-D-glucose 1-phosphate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycosyl group transfer
-
-
-
-
phospho group transfer
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
trehalose degradation V
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-
SYSTEMATIC NAME
IUBMB Comments
alpha,alpha-trehalose:phosphate alpha-D-glucosyltransferase
Unlike EC 2.4.1.64, alpha,alpha-trehalose phosphorylase, this enzyme retains its anomeric configuration. Vanadate is a strong competitive inhibitor of this reversible reaction.
CAS REGISTRY NUMBER
COMMENTARY hide
37205-59-7
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain Horst U1
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-
Manually annotated by BRENDA team
strain Horst U1
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
SwissProt
Manually annotated by BRENDA team
-
-
-
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
2-dehydro-D-glucose + alpha-D-glucose 1-phosphate
? + phosphate
show the reaction diagram
-
-
-
-
?
2-deoxy-D-glucose + alpha-D-glucose 1-phosphate
?
show the reaction diagram
-
8% of the activity with alpha-D-glucose
-
-
r
2-deoxy-D-glucose + alpha-D-glucose 1-phosphate
? + phosphate
show the reaction diagram
-
-
-
-
?
2-fluoro-D-glucose + alpha-D-glucose 1-phosphate
?
show the reaction diagram
-
15% of the activity with alpha-D-glucose
-
-
r
2-fluoro-D-glucose + alpha-D-glucose 1-phosphate
? + phosphate
show the reaction diagram
-
-
-
-
?
2-keto-D-glucose + alpha-D-glucose 1-phosphate
?
show the reaction diagram
-
50% of the activity with alpha-D-glucose
-
-
r
alpha,alpha-trehalose + arsenate
alpha-D-glucose + alpha-D-glucose 1-arsenate
show the reaction diagram
-
-
-
-
r
alpha,alpha-trehalose + arsenate
alpha-D-glucose 1-arsenate + alpha-D-glucose
show the reaction diagram
alpha,alpha-trehalose + phosphate
alpha-D-glucose + alpha-D-glucose 1-phosphate
show the reaction diagram
alpha-D-glucose + alpha-D-glucose 1-phosphate
alpha,alpha-trehalose + phosphate
show the reaction diagram
alpha-D-glucose 1-fluoride + phosphate
alpha-D-glucose 1-phosphate + fluoride
show the reaction diagram
D-mannose + alpha-D-glucose 1-phosphate
?
show the reaction diagram
-
3% of the activity with alpha-D-glucose
-
-
r
D-mannose + alpha-D-glucose 1-phosphate
? + phosphate
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
alpha,alpha-trehalose + phosphate
alpha-D-glucose + alpha-D-glucose 1-phosphate
show the reaction diagram
additional information
?
-
-
the enzyme is involved in trehalose catabolism
-
-
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Mg2+
-
each molecule contains one Mg2+
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1,5-Anhydro-D-glucitol
-
-
1-deoxynojirimycin
2-amino-2-deoxy-D-glucose
-
competitive with glucose
2-deoxy-2-fluoro-alpha-D-glucose 1-fluoride
-
inhibits reaction with alpha,alpha-trehalose
2-deoxy-2-fluoro-glucose 1-phosphate
-
-
ADP
-
34% inhibition at 10 mM
alpha-D-galactose 1-phosphate
-
-
alpha-D-glucose
-
-
alpha-D-glucose 1-fluoride
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competitive with respect to alpha,alpha-trehalose in the phosphorolysis direction
alpha-D-glucose 1-phosphate
-
-
alpha-D-glucuronic acid 1-phosphate
-
-
alpha-D-mannose 1-phosphate
-
-
alpha-D-xylose 1-phosphate
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-
ATP
-
inhibits trehalose degradation, but stimulates synthetic activity
CoCl2
-
39% inhibition at 1 mM
Cu2+
-
complete inactivation at 1 mM
CuSO4
-
37% inhibition at 1 mM
D-galactose
-
-
D-glucono-delta-lactone
-
-
isofagomine
-
-
NaCl
-
competitive with respect to phosphate
orthovanadate
-
competitive against phosphate and alpha-D-glucopyranosyl phosphate, inhibits forward and reverse reaction
phosphate
S-trehalose
-
-
validamycin A
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99% inhibition at 10 mM
validoxylamine A
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vanadate
Zn2+
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inactivation at 1 mM
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
AMP
-
stimulates phosphorolytic activity at 10 mM
ATP
-
stimulates synthetic activity at 10 mM
guanidine
-
no effect on wild-type but guanidine causes a 45fold enhancement of mutant R507A activity
propargylamine
-
no effect on wild-type but propagylamine causes a 23fold enhancement of mutant K512A activity
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
22 - 91
alpha,alpha-trehalose
0.63 - 505
alpha-D-glucose
5.9
alpha-D-glucose 1-fluoride
-
30C, pH 6.6
1.6 - 47
alpha-D-glucose 1-phosphate
40
D-glucose
-
-
0.8 - 103
phosphate
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
4.5
2-dehydro-D-glucose
Pleurotus ostreatus
-
-
0.27
2-deoxy-D-glucose
Pleurotus ostreatus
-
-
0.89
2-fluoro-D-glucose
Pleurotus ostreatus
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-
0.33 - 18.3
alpha,alpha-trehalose
9.5 - 16.8
alpha-D-glucose
0.11
alpha-D-glucose 1-fluoride
Schizophyllum commune
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30C, pH 6.6
9.5 - 16
alpha-D-glucose 1-phosphate
1.15
arsenate
Pleurotus ostreatus
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-
16.8
D-glucose
Pleurotus ostreatus
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-
0.42
D-mannose
Pleurotus ostreatus
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-
13.3 - 17.6
phosphate
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0062 - 0.12
alpha,alpha-trehalose
589
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
46
2-amino-2-deoxy-D-glucose
-
30C, pH 6.6
10.6
2-deoxy-2-fluoro-glucose 1-phosphate
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30C, pH 6.6
11.5
alpha-D-galactose 1-phosphate
-
30C, pH 6.6
0.3
alpha-D-glucose
-
30C, pH 6.6
2.4 - 5.9
alpha-D-glucose 1-phosphate
13.4
alpha-D-glucuronic acid 1-phosphate
-
30C, pH 6.6
1.6
alpha-D-mannose 1-phosphate
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30C, pH 6.6
11
alpha-D-xylose 1-phosphate
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30C, pH 6.6
1000
D-galactose
-
30C, pH 6.6
1000
D-xylose
-
30C, pH 6.6
67
NaCl
-
30C, pH 6.6
1
orthovanadate
-
30C, pH 6.6
2 - 26
phosphate
0.0017
validoxylamine A
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0.0004 - 0.298
vanadate
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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-
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.8 - 6.4
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synthesis of alpha,alpha-trehalose
6 - 7
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synthesis of trehalose
6 - 7.5
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phosphorolytic reaction
6.5 - 6.8
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-
6.7
phosphorolytic reaction
6.8 - 7.2
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phosphorolysis
6.8
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alpha, alpha-trehalose phosphorolysis; assay at
7
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phosphorolytic reaction; synthesis of trehalose
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.5 - 6.8
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pH 5.5: about 80% of maximal activity, pH 75% of maximal activity, synthesis of alpha,alpha-trehalose
5.5 - 7.8
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pH 5.5: about 75% of maximal activity, pH 7.8: about 80% of maxuimal activity, phosphorolysis
6.2 - 6.6
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decrease in enzyme activity below pH 6.2 and above pH 6.6
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
32.5 - 35
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phosphorolysis
35 - 37.5
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synthesis of trehalose
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4.7 - 4.8
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isoelectric focusing between pH 3-9 and pH 3-10
4.8
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isoelectric focusing between pH 4 and pH 6.5
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
55000
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SDS PAGE
58300
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gel filtration
120000
240000
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native PAGE, gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
tetramer
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3.2 - 8.9
-
-
720962
6 - 7
stable
644901
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4
-
unstable, loses activity within a few hours, glycerol, PEG 4000 or trehalose stabilize
20
-
unstable, loses activity within a few hours, glycerol, PEG 4000 or trehalose stabilize
35
-
stable up to 35C, decrease of activity above
additional information
-
PoTPase has a half life of approximately 1 h at 30C
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
highly unstable during purification, glycerol, phosphate, dithiothreitol and PMSF stabilize
-
unstable protein, half-life of no more than approximately 1.3 h in 20mM MES buffer, 25C, pH 7.0. The enzyme can be stabilized on addition of alpha,alpha-trehalose (300 mM) which yields a half-life of 11.5 h. In the presence of 20% (w/v) glycerol, and 9 and 26% (w/v) PEG-4000, half-life is 6.5 h, 16 h and 70 h, respectively. Large improvements in stability by covalent protein modification with activated derivatives of PEG-5000
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-80C, stable for at least 6 months
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
; trehalose phosphorylase from the basidiomycete Pleurotus ostreatus (PoTPase) is isolated from fungal fruit bodies through 500-fold purification with a yield of 44%
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homogeneity
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recombinant enzyme
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D379N
ratio of phosphorolysis to hydrolysis decreases by a factor of 287500
H403A
ratio of phosphorolysis to hydrolysis decreases by a factor of 2190
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
synthesis
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sucrose is converted to alpha,alpha-trehalose in about 60% yield using a coupled enzyme system composed of sucrose phosphorylase from Leuconostoc mesenteroides, glucose isomerase from Streptomyces murinus and the appropriately stabilized PoTPase