Information on EC 2.4.1.245 - alpha,alpha-trehalose synthase

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

EC NUMBER
COMMENTARY hide
2.4.1.245
-
RECOMMENDED NAME
GeneOntology No.
alpha,alpha-trehalose synthase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
NDP-alpha-D-glucose + D-glucose = alpha,alpha-trehalose + NDP
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycosyl group transfer
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-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Metabolic pathways
-
-
Starch and sucrose metabolism
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trehalose biosynthesis VI
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metabolism of disaccharids
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-
SYSTEMATIC NAME
IUBMB Comments
NDP-alpha-D-glucose:D-glucose 1-alpha-D-glucosyltransferase
Requires Mg2+ for maximal activity [1]. The enzyme-catalysed reaction is reversible [1]. In the reverse direction to that shown above, the enzyme is specific for alpha,alpha-trehalose as substrate, as it cannot use alpha- or beta-paranitrophenyl glucosides, maltose, sucrose, lactose or cellobiose [1]. While the enzymes from the thermophilic bacterium Rubrobacter xylanophilus and the hyperthermophilic archaeon Pyrococcus horikoshii can use ADP-, UDP- and GDP-alpha-D-glucose to the same extent [2,3], that from the hyperthermophilic archaeon Thermococcus litoralis has a marked preference for ADP-alpha-D-glucose [1] and that from the hyperthermophilic archaeon Thermoproteus tenax has a marked preference for UDP-alpha-D-glucose [4].
CAS REGISTRY NUMBER
COMMENTARY hide
126341-88-6
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain PRD-1T (DSM 9941)
SwissProt
Manually annotated by BRENDA team
strain PRD-1T (DSM 9941)
SwissProt
Manually annotated by BRENDA team
induced by trehalose and maltose in growth medium
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-
Manually annotated by BRENDA team
strain Kra1(DSM 2078)
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-
Manually annotated by BRENDA team
strain Kra1(DSM 2078)
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-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
the acceptor binding site of TreT shows a wide and commodious groove and lacks the long flexible loop that plays a gating role in ligand binding in trehalose phosphate synthase, TPS, active site, and donor and acceptor binding pocket structure, overview. A wide space at the fissure between two domains and the relative shift of the N-domain in one of the crystal forms suggest that an interactive conformational change between two domains would occur, allowing a more compact architecture for catalysis
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ADP-alpha-D-glucose + D-glucose
ADP + alpha,alpha-1,1-trehalose
show the reaction diagram
ADP-alpha-D-glucose + D-glucose
ADP + alpha,alpha-trehalose
show the reaction diagram
-
-
-
r
ADP-alpha-D-glucose + D-glucose
alpha,alpha-trehalose + ADP
show the reaction diagram
-
GDP-alpha-D-glucose is the most favored in terms of reaction specificity, kcat/Km. UDP-alpha-D-glucose and ADP-alpha-D-glucose are employed with less preferences. The enzyme reversely cleaves alpha,alpha-trehalose to transfer the glucosyl moiety to various NDPs, efficiently producing NDP-alpha-D-glucose. Although ADP-alpha-D-glucose is the least favorable donor, the counterpart, ADP, is the most favorable acceptor for the reverse synthesis of NDP-alpha-D-glucose in kcat/Km. GDP and UDP are less preferred, compared to ADP
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-
r
alpha,alpha-1,1-trehalose + ADP
ADP-glucose + D-glucose
show the reaction diagram
-
-
-
-
r
alpha,alpha-trehalose + ADP
ADP-alpha-D-glucose + D-glucose
show the reaction diagram
-
the enzyme reversely cleaves alpha,alpha-trehalose to transfer the glucosyl moiety to various NDPs, efficiently producing NDP-alpha-D-glucose. Although ADP-alpha-D-glucose is the least favorable donor, the counterpart, ADP, is the most favorable acceptor for the reverse synthesis of NDP-alpha-D-glucose in kcat/Km. GDP and UDP are less preferred, compared to ADP
-
-
r
alpha,alpha-trehalose + GDP
GDP-alpha-D-glucose + D-glucose
show the reaction diagram
-
the enzyme reversely cleaves alpha,alpha-trehalose to transfer the glucosyl moiety to various NDPs, efficiently producing NDP-alpha-D-glucose. Although ADP-alpha-D-glucose is the least favorable donor, the counterpart, ADP, is the most favorable acceptor for the reverse synthesis of NDP-alpha-D-glucose in kcat/Km. GDP and UDP are less preferred, compared to ADP
-
-
r
alpha,alpha-trehalose + UDP
UDP-alpha-D-glucose + D-glucose
show the reaction diagram
-
the enzyme reversely cleaves alpha,alpha-trehalose to transfer the glucosyl moiety to various NDPs, efficiently producing NDP-alpha-D-glucose. Although ADP-alpha-D-glucose is the least favorable donor, the counterpart, ADP, is the most favorable acceptor for the reverse synthesis of NDP-alpha-D-glucose in kcat/Km. GDP and UDP are less preferred, compared to ADP
-
-
r
GDP-alpha-D-glucose + D-glucose
alpha,alpha-trehalose + GDP
show the reaction diagram
-
GDP-alpha-D-glucose is the most favored in terms of reaction specificity, kcat/Km. UDP-alpha-D-glucose and ADP-alpha-D-glucose are employed with less preferences. The enzyme reversely cleaves alpha,alpha-trehalose to transfer the glucosyl moiety to various NDPs, efficiently producing NDP-alpha-D-glucose. Although ADP-alpha-D-glucose is the least favorable donor, the counterpart, ADP, is the most favorable acceptor for the reverse synthesis of NDP-alpha-D-glucose in kcat/Km. GDP and UDP are less preferred, compared to ADP
-
-
r
GDP-glucose + D-glucose
alpha,alpha-1,1-trehalose + GDP
show the reaction diagram
GDP-glucose + D-glucose
alpha,alpha-trehalose + GDP
show the reaction diagram
-
-
-
r
UDP-alpha-D-glucose + D-glucose
alpha,alpha-trehalose + UDP
show the reaction diagram
-
GDP-alpha-D-glucose is the most favored in terms of reaction specificity, kcat/Km. UDP-alpha-D-glucose and ADP-alpha-D-glucose are employed with less preferences. The enzyme reversely cleaves alpha,alpha-trehalose to transfer the glucosyl moiety to various NDPs, efficiently producing NDP-alpha-D-glucose. Although ADP-alpha-D-glucose is the least favorable donor, the counterpart, ADP, is the most favorable acceptor for the reverse synthesis of NDP-alpha-D-glucose in kcat/Km. GDP and UDP are less preferred, compared to ADP
-
-
r
UDP-glucose + D-glucose
alpha,alpha-1,1-trehalose + UDP
show the reaction diagram
UDP-glucose + D-glucose
alpha,alpha-trehalose + UDP
show the reaction diagram
-
-
-
r
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
ADP-alpha-D-glucose + D-glucose
ADP + alpha,alpha-trehalose
show the reaction diagram
O58762
-
-
-
r
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Co2+
-
72% of the activity with Mg2+
Fe3+
2 mM Fe3+ induces slightly enhanced activity
Li+
2 mM or 20 mM Li+ induces slightly enhanced activity
Ni2+
-
57% of the activity with Mg2+
additional information
no enhanced activity in the presence of 20 mM Fe3+ or Mn2+
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
6.8 - 14.19
ADP
3.2
ADP-alpha-D-glucose
-
pH 6.0, 60°C
0.8 - 1.14
ADP-glucose
11.5 - 82
alpha,alpha-1,1-trehalose
25.24
alpha,alpha-trehalose
-
pH 6.0, 60°C
1.3 - 6.2
D-glucose
23.77
GDP
-
pH 6.0, 60°C
2.08
GDP-alpha-D-glucose
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pH 6.0, 60°C
29.54
UDP
-
pH 6.0, 60°C
2.46
UDP-alpha-D-glucose
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pH 6.0, 60°C
0.23
UDP-glucose
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in 50 mM HEPES/KOH (pH 7.0), with 20 mM MgCl2, at 70°C
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.7
ADP
Pyrococcus horikoshii
-
pH 6.0, 60°C
0.44
ADP-alpha-D-glucose
Pyrococcus horikoshii
-
pH 6.0, 60°C
1.2
alpha,alpha-trehalose
Pyrococcus horikoshii
-
pH 6.0, 60°C
1.7
GDP
Pyrococcus horikoshii
-
pH 6.0, 60°C
0.54
GDP-alpha-D-glucose
Pyrococcus horikoshii
-
pH 6.0, 60°C
1.5
UDP
Pyrococcus horikoshii
-
pH 6.0, 60°C
0.53
UDP-alpha-D-glucose
Pyrococcus horikoshii
-
pH 6.0, 60°C
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.12
ADP
Pyrococcus horikoshii
-
pH 6.0, 60°C
13
0.14
ADP-alpha-D-glucose
Pyrococcus horikoshii
-
pH 6.0, 60°C
11248
0.05
alpha,alpha-trehalose
Pyrococcus horikoshii
-
pH 6.0, 60°C
589
0.07
GDP
Pyrococcus horikoshii
-
pH 6.0, 60°C
53
0.26
GDP-alpha-D-glucose
Pyrococcus horikoshii
-
pH 6.0, 60°C
3886
0.05
UDP
Pyrococcus horikoshii
-
pH 6.0, 60°C
26
0.22
UDP-alpha-D-glucose
Pyrococcus horikoshii
-
pH 6.0, 60°C
364
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.5
-
13% residual activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
100
-
65% residual activity
PDB
SCOP
CATH
ORGANISM
UNIPROT
Pyrococcus horikoshii (strain ATCC 700860 / DSM 12428 / JCM 9974 / NBRC 100139 / OT-3)
Pyrococcus horikoshii (strain ATCC 700860 / DSM 12428 / JCM 9974 / NBRC 100139 / OT-3)
Pyrococcus horikoshii (strain ATCC 700860 / DSM 12428 / JCM 9974 / NBRC 100139 / OT-3)
Pyrococcus horikoshii (strain ATCC 700860 / DSM 12428 / JCM 9974 / NBRC 100139 / OT-3)
Pyrococcus horikoshii (strain ATCC 700860 / DSM 12428 / JCM 9974 / NBRC 100139 / OT-3)
Pyrococcus horikoshii (strain ATCC 700860 / DSM 12428 / JCM 9974 / NBRC 100139 / OT-3)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
44400
-
subunit, calculated from amino acid sequence
46700
calculated from amino acid sequence
47000
-
subunit, SDS-PAGE
48000
-
2 * 48146, calculated, 2 * 48000, SDS-PAGE
48146
-
2 * 48146, calculated, 2 * 48000, SDS-PAGE
48197
-
x * 48197, calculated, x * 49871, MALDI-TOF mass spectrometry of recombinant His-tagged protein
49871
-
x * 48197, calculated, x * 49871, MALDI-TOF mass spectrometry of recombinant His-tagged protein
100000
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native enzyme, gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
-
x * 48197, calculated, x * 49871, MALDI-TOF mass spectrometry of recombinant His-tagged protein
dimer
-
2 * 48146, calculated, 2 * 48000, SDS-PAGE
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified enzyme free or as TreT-UDP binary complex, 10 mg/ml native enzyme from PEG 3350 25%, 0.2 M MgCl2, and 0.1 M sodium HEPES, 18°C, the selenomethionine-substituted protein crystal grow from 21% methoxy PEG 2000, 0.18 M ammonium sulfate, and 0.1 M sodium acetate, pH 4.6, for the UDP-glucose complex crystal, 5 mM UDPG is added to 10 mg/ml E326A protein for 1 h prior to setup of the crystallization using the same conditions as for the native crystal, X-ray diffraction structure determination and analysis at 2.3-3.0 A resolution, single-wavelength anomalous dispersion phasing
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 8
-
highly stable within this range
660897
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
55
-
1 h, 25% residual activity
60 - 70
the half-lives for inactivation at 60°C and at 70°C are 309 h and 4.1 h, respectively
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
HisTrap column chromatography, Q-Sepharose fast-flow column chromatography, and Superdex 200 gel filtration
Ni-NTA affinity column chromatography
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli BL21(DE3) cells
expression in Escherichia coli with His6-tag
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expression in Escherichia coli, with His-tag
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gene treT, DNA and amino acid sequence determination and analysis, sequence comparison
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D134E
the mutant shows 10% reduced activity in the forward reaction, and 50% reduced activity in the reverse reaction
D134R
inactive mutant
D274R/D275A
inactive mutant
E326A
inactive mutant
F85R
the mutant shows 95% reduced activity in the forward reaction, and no activity in the reverse reaction
F85Y
the mutant shows 10% reduced activity in the forward reaction, and 40% reduced activity in the reverse reaction
H155D
inactive in the forward reaction, 80% reduced activity in the reverse reaction compared to wild-type
H92A
the mutant shows wild-type activity in the forward reaction, and 90% reduced activity in the reverse reaction
K209R
the mutant shows wild-type activity
Q96A
the mutant shows 10% reduced activity in the forward reaction, and 90% reduced activity in the reverse reaction
R239A
inactive in the reverse reaction, 10% reduced activity in the forward reaction compared to wild-type
T49H
the mutant shows 90% reduced activity in the forward reaction, and 98% reduced activity in the reverse reaction
T49R
inactive mutant
V309L
the mutant shows wild-type activity in the forward reaction, and 25% reduced activity in the reverse reaction
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
synthesis