Information on EC 2.7.1.175 - maltokinase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
2.7.1.175
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RECOMMENDED NAME
GeneOntology No.
maltokinase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + maltose = ADP + alpha-maltose 1-phosphate
show the reaction diagram
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Metabolic pathways
-
-
Starch and sucrose metabolism
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SYSTEMATIC NAME
IUBMB Comments
ATP:alpha-maltose 1-phosphotransferase
Requires Mg2+ for activity.
CAS REGISTRY NUMBER
COMMENTARY hide
179987-43-0
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GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
metabolism
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
ATP + maltose
ADP + alpha-maltose 1-phosphate
show the reaction diagram
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-
product analysis and determination by chemical analysis and NMR spectroscopy
-
?
ATP + maltose
ADP + alpha-maltose-1-phosphate
show the reaction diagram
ATP + maltose
ADP + maltose 1-phosphate
show the reaction diagram
GTP + maltose
GDP + maltose 1-phosphate
show the reaction diagram
-
-
NMR spectroscopical product analysis
-
?
UTP + maltose
UDP + maltose 1-phosphate
show the reaction diagram
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NMR spectroscopical product analysis
-
?
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
ATP + maltose
ADP + alpha-maltose 1-phosphate
show the reaction diagram
-
-
-
-
?
ATP + maltose
ADP + alpha-maltose-1-phosphate
show the reaction diagram
ATP + maltose
ADP + maltose 1-phosphate
show the reaction diagram
-
-
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
GTP
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can substitute for ATP
UTP
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can substitute for ATP
additional information
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CTP and TTP show no activity, while GTP with 2.1% activity compared to ATP and UTP with 2.7% show only poor activity as phosphoryl-group donors
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Co2+
-
activates, 41% activity compared to Mg2+
Mn2+
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activates, 14% activity compared to Mg2+
NaCl
-
addition of 50 mM NaCl markedly enhances Mak stability
additional information
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divalent cations are required for activity and Mg2+ is the best activator
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
glycerol
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strongly inhibits Mak activity, even at very low concentrations of 1%
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
Pep2 forms a heterooctameric complex with trehalose synthase TreS, the complex formation markedly accelerates the maltokinase activity of Pep2, overview
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.54 - 0.74
ATP
2.52 - 2.6
maltose
additional information
additional information
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
144.8
purified His-tagged recombinant enzyme, pH 7.0, 45°C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7 - 9
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at 37°C
7.75
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in Tris/HCl buffer
7.8 - 8
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assay at
8.5
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in TEA/HCl buffer
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 11
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activity range
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20 - 65
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activity range
additional information
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the activity of maltokinase increases 2.5fold with a maximum at 55°C, but decreases rapidly at temperatures above 60°C
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4.3
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isoelectric focusing
PDB
SCOP
CATH
ORGANISM
UNIPROT
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium vanbaalenii (strain DSM 7251 / PYR-1)
Mycobacterium vanbaalenii (strain DSM 7251 / PYR-1)
Mycobacterium vanbaalenii (strain DSM 7251 / PYR-1)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
49800
x * 49800, His-tagged enzyme, sequence calculation, x * 62000, recombinant His-tagged enzyme, SDS-PAGE
50700
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1 * 50700, recombinant His-tagged Mak, SDS-PAGE
52400
x * 52400, His-tagged enzyme, sequence calculation, x * 53000, recombinant His-tagged enzyme, SDS-PAGE
53000
x * 52400, His-tagged enzyme, sequence calculation, x * 53000, recombinant His-tagged enzyme, SDS-PAGE
57000
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1 * 57000, SDS-PAGE
62000
x * 49800, His-tagged enzyme, sequence calculation, x * 62000, recombinant His-tagged enzyme, SDS-PAGE
452900
TreS-Pep2 complex, sequence calculation
470000
TreS-Pep2 complex, ultracentrifugation, Pep2 forms a heterooctameric complex with trehalose synthase TreS, the complex formation markedly accelerates the maltokinase activity of Pep2
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
trimer or tetramer
additional information
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified recombinant enzyme in complex with a non-hydrolysable ATP analogue, mixing of 0.001 ml of protein in 20 mg/ml in 50 mM BTP, pH 7.5, 50 mM NaCl, with 0.0007-0.001 ml of reservoir solution containing 0.1 M MOPS/sodium HEPES, pH 7.5, 0.12 M ethylene glycols (0.03 M each of di-, tri-, tetra-, and penta-ethyleneglycol), 30% PEG 500 MME/PEG 20000, and equilibration against 00.3 ml reservoir solution, 20°C, microseeding, X-ray diffraction structure determination and analysis at 1.15 A resolution, modelling. Crystallzation attempts of the enzyme in complex with maltose are all unsuccessful
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
addition of 50 mM NaCl markedly enhances Mak stability
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
4°C, purified recombinant His-tagged enzyme, 7 days, about 40% of activity remaining, at pH 7.5, most of the activity is lost after freeze/thawing at -20°C. 10 mM maltose only slightly improve the stability at 4°C to about 50% of maximal activity, but addition of 50 mM NaCl dramatically improve the stability of the enzyme with over 90% activity remaining after 1 week at 4°C
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
native enzyme by heat treatment at 50°C for 30 min, anion exchange chromatography, ammonium sulfate fractionation, hydrophobic interaction chromatography, ultra- and gel filtration, and another step of anion exchange chromatography
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recombinant C-terminally His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity and anion exchange chromatography, followed by gel filtration
recombinant enzyme from Escherichia coli by anion exchange chromatography and gel filtration
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recombinant His-tagged enzyme from Mycobacterium smegmatis by nickel affinity chromatography
recombinant His-tagged enzyme from Streptomyces lividans strain 66 by nickel affinity chromatography
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
gene mak, DNA and amino acid sequence determination and analysis, high level overexpression of His-tagged Mak in Escherichia coli
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gene mak, DNA and amino acid sequence determination and analysis, phylogenetic analysis, recombinant expression of C-terminally His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
gene mak, phylogenetic analysis, recombinant expression of C-terminally His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
gene mak1, located in a gene cluster, DNA and amino acid sequence determination and analysis, sequence comparisons, expression of the His-tagged enzyme in Streptomyces lividans strain 66 using vector pMJP7
gene pep2, sequence comparisons, expression of the His-tagged enzyme in Streptomyces lividans strain 66 using vector pMJP7
gene Rv0127 or pep2, recombinant overexpression of His-tagged enzyme in Mycobacterium smegmatis
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D321A
site-directed mutagenesis, the Mg2+ binding residue mutant is inactive
D339N
inactive mutant
E340R
site-directed mutagenesis, the mutant shows decreased activity compared to the wild-type enzyme
K145A
site-directed mutagenesis, the Mg2+ binding residue mutant is inactive
N145A
site-directed mutagenesis, inactive mutant
Q309A
site-directed mutagenesis, the Mg2+ binding residue mutant is inactive
R351A
site-directed mutagenesis, the mutant shows decreased activity compared to the wild-type enzyme
S144A
site-directed mutagenesis, the mutant shows highly decreased activity compared to the wild-type enzyme
D321A
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site-directed mutagenesis, the Mg2+ binding residue mutant is inactive
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D339N
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inactive mutant
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E340R
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site-directed mutagenesis, the mutant shows decreased activity compared to the wild-type enzyme
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K145A
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site-directed mutagenesis, the Mg2+ binding residue mutant is inactive
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N145A
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site-directed mutagenesis, inactive mutant
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Q309A
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site-directed mutagenesis, the Mg2+ binding residue mutant is inactive
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R351A
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site-directed mutagenesis, the mutant shows decreased activity compared to the wild-type enzyme
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S144A
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site-directed mutagenesis, the mutant shows highly decreased activity compared to the wild-type enzyme
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D334N
inactive mutant
E324R
site-directed mutagenesis, the mutant shows decreased activity compared to the wild-type enzyme
K413A
site-directed mutagenesis
N137A
site-directed mutagenesis, inactive mutant
R334R
site-directed mutagenesis, the mutant shows decreased activity compared to the wild-type enzyme
S136A
site-directed mutagenesis, the mutant shows highly decreased activity compared to the wild-type enzyme
Y416A
site-directed mutagenesis
Y416F
site-directed mutagenesis
Y420A
site-directed mutagenesis
Y420F
site-directed mutagenesis
N137A
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site-directed mutagenesis, inactive mutant
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S136A
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site-directed mutagenesis, the mutant shows highly decreased activity compared to the wild-type enzyme
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Y416A
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site-directed mutagenesis
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Y420A
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site-directed mutagenesis
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Y420F
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site-directed mutagenesis
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
drug development
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Mak may be a potential drug target in Mycobacterium tuberculosis
synthesis