Information on EC 5.4.99.15 - (1->4)-alpha-D-Glucan 1-alpha-D-glucosylmutase

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

EC NUMBER
COMMENTARY hide
5.4.99.15
-
RECOMMENDED NAME
GeneOntology No.
(1->4)-alpha-D-Glucan 1-alpha-D-glucosylmutase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
4-[(1->4)-alpha-D-glucosyl]n-1-D-glucose = 1-alpha-D-[(1->4)-alpha-D-glucosyl]n-1-alpha-D-glucopyranoside
show the reaction diagram
-
-
-
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
intramolecular transglucosylation
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Biosynthesis of secondary metabolites
-
-
Metabolic pathways
-
-
Starch and sucrose metabolism
-
-
trehalose biosynthesis V
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metabolism of disaccharids
-
-
SYSTEMATIC NAME
IUBMB Comments
(1->4)-alpha-D-glucan 1-alpha-D-glucosylmutase
The enzyme from Arthrobacter sp., Sulfolobus acidocaldarius acts on (1->4)-alpha-D-glucans containing three or more (1->4)-alpha-linked D-glucose units. Not active towards maltose.
CAS REGISTRY NUMBER
COMMENTARY hide
170780-49-1
-
171265-12-6
cellulase (Rhizobium strain M11 clone pBMT7 trehalose oligosaccharide-forming) /maltooligosyl trehalose synthase (Rhizobium strain M-11 clone pBMTU1 gene treY reduced) /synthase, maltooligosyltrehalose (Rhizobium strain M-11 clone pBMTU1 gene treY reduced)
171265-13-7
cellulase (Arthrobacter strain Q36 clone pBMT13 trehalose oligosaccharide-forming) /synthase, maltooligosyltrehalose (Arthrobacter clone pBRT4 gene treY)
184890-98-0
synthase, maltooligosyltrehalose (Sulfolobus acidocaldarius clone pBST1 gene treY)
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain SN223/29
-
-
Manually annotated by BRENDA team
strain SN223/29
-
-
Manually annotated by BRENDA team
strain 7120
-
-
Manually annotated by BRENDA team
strain 7120
-
-
Manually annotated by BRENDA team
Q36
-
-
Manually annotated by BRENDA team
Q36
-
-
Manually annotated by BRENDA team
presence of three enzymes for trehalose synthesis: trehalose synthase, maltooligosyltrehalose synthase, and trehalose-6-phosphate synthetase
-
-
Manually annotated by BRENDA team
presence of three enzymes for trehalose synthesis: trehalose synthase, maltooligosyltrehalose synthase, and trehalose 6-phosphate synthetase
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-
Manually annotated by BRENDA team
strain JCM8857
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-
Manually annotated by BRENDA team
M-11
-
-
Manually annotated by BRENDA team
M-11
-
-
Manually annotated by BRENDA team
strain MT4
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-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
physiological function
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
acarbose
?
show the reaction diagram
alpha-maltotriosyl trehalose
maltopentaose
show the reaction diagram
-
-
-
?
dextrin
?
show the reaction diagram
liquified corn starch
?
show the reaction diagram
Maltoheptaose
?
show the reaction diagram
maltoheptaose
alpha-maltopentaosyl trehalose
show the reaction diagram
Maltohexaose
?
show the reaction diagram
maltohexaose
alpha-maltotetraosyl trehalose
show the reaction diagram
maltohexaose
alpha-maltotetraosyltrehalose
show the reaction diagram
maltohexaose
trehalosylmaltotetraose
show the reaction diagram
maltopentaose
?
show the reaction diagram
Maltopentaose
alpha-Maltotriosyl trehalose
show the reaction diagram
maltopentaose
alpha-maltotriosyltrehalose
show the reaction diagram
Maltotetraose
?
show the reaction diagram
maltotetraose
alpha-maltosyl trehalose
show the reaction diagram
-
ratio of transglycosylation to hydrolysis is 38:0.6, recombinant enzyme
i.e. alpha-D-Glcp-(1->4)-alpha-D-Glcp-(1->4)-alpha-D-Glcp-(1->1)-alpha-D-Glcp
-
?
maltotetraose
alpha-maltosyltrehalose
show the reaction diagram
maltotriose
?
show the reaction diagram
-
-
-
?
maltotriose
alpha-glucosyltrehalose
show the reaction diagram
Short chain amylose
?
show the reaction diagram
-
43% of the activity with maltopentaose
-
-
-
soluble starch
?
show the reaction diagram
soluble starch
trehalose
show the reaction diagram
-
-
-
-
?
starch
?
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
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ca2+
EF433294
134% relative activity at 1 mM, at 50°C in 50 mM phosphate buffer (pH 7.0)
Na+
-
triggers maximum expression and activity of MTSase
Zn2+
EF433294
122% relative activity at 1 mM, at 50°C in 50 mM phosphate buffer (pH 7.0)
additional information
-
metal ions are not required for activity. Ca2+, K+, Mg2+, or Na+ have no activating or inhibiting effect on activity
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
EDTA
EF433294
31% residual activity at 1 mM, at 50°C in 50 mM phosphate buffer (pH 7.0)
Mg2+
EF433294
66% residual activity at 1 mM, at 50°C in 50 mM phosphate buffer (pH 7.0)
MgCl2
-
weak
Mn2+
EF433294
51% residual activity at 1 mM, at 50°C in 50 mM phosphate buffer (pH 7.0)
validamycin A
Zn2+
-
4 mM, about 60% loss of activity
ZnSO4
-
weak
additional information
-
Ca2+, K+, Mg2+, or Na+ have no activating or inhibiting effect on activity
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
NaCl
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exposure to 150 mM NaCl for 48 h causes increased expression of MTSase
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.9 - 5.71
maltoheptaose
1.4 - 26.5
maltohexaose
5 - 122
maltopentaose
11.6 - 41.5
maltotetraose
228
maltotriose
-
pH 5.5, 60°C
0.6
short chain amylose
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-
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
147 - 429
maltoheptaose
6.11 - 445
maltohexaose
13.4 - 530
maltopentaose
61 - 196
maltotetraose
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.31 - 62.3
maltopentaose
272
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
14.4
-
recombinant enzyme, after 45.5fold purification, at 70°C
19
-
pH 5.0, 75°C, substrate: maltohexaose
550
-
pH 5.5, 75°C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 6
-
MTSH fusion enzyme
7.5
-
in 100 mM phosphate buffer
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4.3 - 6
-
pH 4.3: about 75% of maximal activity, pH 6.0: about 80% of maximal activity
5.5 - 8
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pH 5.5: about 80% of maximal activity, pH 8.0: about 60% of maximal activity
6 - 8
EF433294
-
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50
EF433294
recombinant enzyme
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 45
-
30°C: about 80% of maximal activity, 45°C: about 85% of maximal activity
45 - 90
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45°C: about 60% of maximal activity, 90°C: about 55% of maximal activity
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5
-
FPLC chromatofocusing
6.1
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isoelectric focusing
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
additional information
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
70000
-
x * 70000, about, sequence calculation
76000
-
x * 76000, SDS-PAGE
80000
-
gel filtration
81000
-
x * 81000, SDS-PAGE
83800
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x * 83800, calculated, x * 90000, SDS-PAGE
86609
-
x * 86609, calculated from sequence
90000
-
x * 83800, calculated, x * 90000, SDS-PAGE
106600
EF433294
calculated from amino acid sequence
114000
-
SDS-PAGE
125000
EF433294
His6-tagged enzyme, SDS-PAGE
150000
-
MTSH fusion enzyme
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
hanging-drop vapour-diffusion method at 4°C. The chemical modification of this enzyme by reductive methylation of lysine residues significantly improves the crystal quality for X-ray diffraction experiments. The crystals of the modified enzyme belong to orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 56.70, b = 140.1, c = 205.2 A measured at cryo-temperature, and are found to contain two enzyme molecules per asymmetric unit; recombinant enzyme expressed in Escherichia coli, hanging-drop vapour-diffusion method, chemical modification of this enzyme by reductive methylation of lysine residues significantly improves the crystal quality for X-ray diffraction experiments
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hanging drop vapor diffusion method, using 0.2 M potassium sodium tartrate tetrahydrate, 20% (w/v) polyethylene glycol 3350
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4 - 10
-
stable
721858, 722996
4.5 - 9.5
4.5 - 11
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24 h, stable
724948
6 - 9.5
-
4°C, 24 h, stable
3486
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
40
-
60 min, stable up to
70 - 80
-
the MTSH fusion enzyme exhibits a high degree of thermostability, retaining 80% of the activity when pre-incubated at 70°C for 48 h, the stability is gradually abolished by incubating the fusion enzyme at above 80°C for 30 min
70
-
48 h, 80% residual activity
75
-
MTS proves stable at 75°C for more than 24 h, with a half life of about 50 h
90
-
1 h, 30% loss of activity
95
-
2 h, 90% loss of activity
100
-
1 h, 65% loss of activity
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
; recombinant enzyme expressed in Escherichia coli
-
gel filtration
-
Ni-NTA column chromatography
-
Ni2+-NTA-agarose column chromatography
EF433294
recombinant wild-type and mutants
-
Resource ISO column chromatography, Resource Q column chromatography, and Superdex 200 gel filtration
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cloned from the genomic DNA of Sulfolobus solfataricus ATCC 35092 to an expression vector with a T7 lac promoter and then expressed in Escherichia coli
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expressed in Escherichia coli BL21 cells
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expressed in Escherichia coli BL21(DE3) cells
EF433294
expressed in Escherichia coli BL21(DE3)pLysS cells
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expressed in Escherichia coli BL21(DE3)pLysS cells as fusion enzyme with maltooligosyltrehalose trehalohydrolase
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expressed in Lactococcus lactis strain NZ9000
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expression in Escherichia coli
gene mts, DNA and amino acid sequence determination and analysis, expression analysis
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inserted into an expression vector, pBV220, to yield the recombinant plasmid pSBGT1. MTSase gene in pBSGT1 is expressed in Escherichia coli
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overexpression in Escherichia coli using the T7-expression system
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overexpression of enzyme and maltooligosyltrehalose trehalohydrolase alone or with UPD-glucose pyrophosphorylase in a synthetic operon to improve trehalose production in Escherichia coli
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the gene of maltooligosyltrehalose synthase is amplified by PCR. The primers are designed according to the published sequence of homologous gene from Sulfolobus acidocaldarius ATCC33909. This gene is inserted into the plasmid pBV220 and the resultant recombinant plasmid pBV220-GT is transformed to Escherichia coli DH5 alpha
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wild-type and mutant enzymes are expressed in Escherichia coli Rosetta (DE3)
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
the expression of MTS is upregulated during drought stress
EF433294
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
up
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enzyme expression, as well as accumulations of both trehalose and especially sucrose in filaments, are upregulated significantly under dehydration stress, NaCl stress, and high temperature-drought stress
D228N
-
complete loss of both hydrolytic and transglycosylation activity
D231A
-
significant decrease of both hydrolytic and transglycosylation activity
D275A
-
about 50% and 40% decrease of hydrolytic and transglycosylation activity, resp.
D394A
-
significant decrease of both hydrolytic and transglycosylation activity
D443N
-
complete loss of both hydrolytic and transglycosylation activity
E255Q
-
complete loss of both hydrolytic and transglycosylation activity
E393A
-
complete loss of both hydrolytic and transglycosylation activity
H229N
-
complete loss of both hydrolytic and transglycosylation activity
K256T
-
significant decrease of both hydrolytic and transglycosylation activity
K390A
-
increase in hydrolytic, decrease in transglycosylation activity
K390E
-
increase in hydrolytic, decrease in transglycosylation activity
K390H
-
increase in hydrolytic, decrease in transglycosylation activity
K390Q
-
increase in hydrolytic, decrease in transglycosylation activity
K390R
-
increase in hydrolytic, decrease in transglycosylation activity
K390T
-
increase in hydrolytic, decrease in transglycosylation activity; increase in hydrolytic, significant decrease in transglycosylation activity
K390W
-
complete loss of transglycosylation activity, increase in hydrolytic activity
K390W/K445W
-
complete loss of transglycosylation activity, 40% decrease in hydrolytic activity
K445A
-
increase in hydrolytic, decrease in transglycosylation activity
K445E
-
increase in hydrolytic, decrease in transglycosylation activity
K445H
-
increase in hydrolytic, decrease in transglycosylation activity
K445Q
-
increase in hydrolytic, decrease in transglycosylation activity
K445R
-
increase in hydrolytic, decrease in transglycosylation activity
K445T
-
increase in hydrolytic, significant decrease in transglycosylation activity
K445W
-
complete loss of transglycosylation activity, increase in hydrolytic activity
R192A
-
about 50% and 40% decrease of hydrolytic and transglycosylation activity, resp.
R402S
-
about 30% decrease of both hydrolytic and transglycosylation activity
Y274S
-
significant decrease of both hydrolytic and transglycosylation activity
Y400S
-
about 25% and 20% decrease of hydrolytic and transglycosylation activity, resp.
A406
-
compared with wild-type enzyme, the hydrolysis:transglycosylation selectivity ratio is significantly increased
A406S
-
the hydrolysis/transglycosylation selectivity ratio shows little change
D411A
-
significant reductions in catalytic efficiency and increase in the transition-state energy; significant reductions in catalytic efficiency, increase in the transition-state energy
D610A
-
significant reductions in catalytic efficiency and increase in the transition-state energy; significant reductions in catalytic efficiency, increase in the transition-state energy
F206W
-
significant decrease in transglycosylation activity
F206Y
-
significant decrease in transglycosylation activity
F207Y
-
significant decrease in transglycosylation activity
F405M
-
catalytic activity comparable to wild-type
F405S
-
depending on substrate, 29-97% of wild-type transglycosylation efficiency, hydrolytic activity is 16% of wild-type
F405W
-
catalytic activity slightly lower than wild-type
P402
-
compared with wild-type enzyme, the hydrolysis:transglycosylation selectivity ratio is significantly decreased
P402Q
-
the hydrolysis/transglycosylation selectivity ratio is significantly decreased
R614A
-
significant reductions in catalytic efficiency and increase in the transition-state energy; significant reductions in catalytic efficiency, increase in the transition-state energy
V426
-
little change in hydrolysis:transglycosylation selectivity ratio
Y290F
-
mutation located near subsite +1 constructed to alter enzyme selectivity. Catalytic efficiency for hydrolysis and transglycosylation reaction are 6.6 and 5.6% resp., of wild-type
Y367F
-
mutation located near subsite +1 constructed to alter enzyme selectivity. Catalytic efficiency for hydrolysis and transglycosylation reaction are about half of wild-type
Y409F
-
mutation located near subsite +1 constructed to alter enzyme selectivity. Catalytic efficiency for hydrolysis is similar to wild-type, for transglycosylation reaction somewhat lower than wild-type
A406S
-
the hydrolysis/transglycosylation selectivity ratio shows little change
-
D411A
-
significant reductions in catalytic efficiency and increase in the transition-state energy
-
D610A
-
significant reductions in catalytic efficiency and increase in the transition-state energy
-
P402Q
-
the hydrolysis/transglycosylation selectivity ratio is significantly decreased
-
R614A
-
significant reductions in catalytic efficiency and increase in the transition-state energy
-
A406
-
compared with wild-type enzyme, the hydrolysis:transglycosylation selectivity ratio is significantly increased
-
D610A
-
significant reductions in catalytic efficiency, increase in the transition-state energy
-
P402
-
compared with wild-type enzyme, the hydrolysis:transglycosylation selectivity ratio is significantly decreased
-
R614A
-
significant reductions in catalytic efficiency, increase in the transition-state energy
-
V426
-
little change in hydrolysis:transglycosylation selectivity ratio
-
additional information
-
overexpression of BvMTSH, a fusion gene for maltooligosyltrehalose synthase and maltooligosyltrehalose trehalohydrolase, enhances drought tolerance in transgenic Oryza sativa without growth inhibition. The transgenic pants show an abscisic acid-hyposensitive phenotype in the roots, phenotype, overview. Construction of the bifunctional in-frame fusion of maltooligosyltrehalose synthase and maltooligosyltrehalose trehalohydrolase from the nonpathogenic bacterium Brevibacterium helvolum under the control of the constitutive rice cytochrome c promoter (101MTSH) or the ABA-inducible Ai promoter (105MTSH). BvMTS converts maltooligosaccharides into maltooligosyltrehalose and BvMTH releases trehalose
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
synthesis
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