BRENDA - Enzyme Database
show all sequences of 2.7.1.175

Structure of mycobacterial maltokinase, the missing link in the essential GlgE-pathway

Fraga, J.; Maranha, A.; Mendes, V.; Pereira, P.J.; Empadinhas, N.; Macedo-Ribeiro, S.; Sci. Rep. 5, 8026 (2015)

Data extracted from this reference:

Cloned(Commentary)
Commentary
Organism
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)
Mycolicibacterium vanbaalenii
gene mak, phylogenetic analysis, recombinant expression of C-terminally His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
Mycobacterium tuberculosis
Crystallization (Commentary)
Crystallization
Organism
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
Mycolicibacterium vanbaalenii
Engineering
Amino acid exchange
Commentary
Organism
D334N
inactive mutant
Mycolicibacterium vanbaalenii
D339N
inactive mutant
Mycobacterium tuberculosis
E324R
site-directed mutagenesis, the mutant shows decreased activity compared to the wild-type enzyme
Mycolicibacterium vanbaalenii
E340R
site-directed mutagenesis, the mutant shows decreased activity compared to the wild-type enzyme
Mycobacterium tuberculosis
K413A
site-directed mutagenesis
Mycolicibacterium vanbaalenii
N137A
site-directed mutagenesis, inactive mutant
Mycolicibacterium vanbaalenii
N145A
site-directed mutagenesis, inactive mutant
Mycobacterium tuberculosis
R334R
site-directed mutagenesis, the mutant shows decreased activity compared to the wild-type enzyme
Mycolicibacterium vanbaalenii
R351A
site-directed mutagenesis, the mutant shows decreased activity compared to the wild-type enzyme
Mycobacterium tuberculosis
S136A
site-directed mutagenesis, the mutant shows highly decreased activity compared to the wild-type enzyme
Mycolicibacterium vanbaalenii
S144A
site-directed mutagenesis, the mutant shows highly decreased activity compared to the wild-type enzyme
Mycobacterium tuberculosis
Y416A
site-directed mutagenesis
Mycolicibacterium vanbaalenii
Y416F
site-directed mutagenesis
Mycolicibacterium vanbaalenii
Y420A
site-directed mutagenesis
Mycolicibacterium vanbaalenii
Y420F
site-directed mutagenesis
Mycolicibacterium vanbaalenii
Metals/Ions
Metals/Ions
Commentary
Organism
Structure
Mg2+
required
Mycobacterium tuberculosis
Mg2+
required, magnesium-binding residue is Asp322
Mycolicibacterium vanbaalenii
Natural Substrates/ Products (Substrates)
Natural Substrates
Organism
Commentary (Nat. Sub.)
Natural Products
Commentary (Nat. Pro.)
Organism (Nat. Pro.)
Reversibility
ATP + maltose
Mycolicibacterium vanbaalenii
-
ADP + alpha-maltose-1-phosphate
-
-
?
ATP + maltose
Mycobacterium tuberculosis
-
ADP + alpha-maltose-1-phosphate
-
-
?
ATP + maltose
Mycolicibacterium vanbaalenii DSM 7251
-
ADP + alpha-maltose-1-phosphate
-
-
?
ATP + maltose
Mycobacterium tuberculosis ATCC 25618
-
ADP + alpha-maltose-1-phosphate
-
-
?
Organism
Organism
Primary Accession No. (UniProt)
Commentary
Textmining
Mycobacterium tuberculosis
O07177
-
-
Mycobacterium tuberculosis ATCC 25618
O07177
-
-
Mycolicibacterium vanbaalenii
A1TH50
-
-
Mycolicibacterium vanbaalenii DSM 7251
A1TH50
-
-
Purification (Commentary)
Commentary
Organism
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
Mycobacterium tuberculosis
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
Mycolicibacterium vanbaalenii
Reaction
Reaction
Commentary
Organism
ATP + maltose = ADP + alpha-maltose 1-phosphate
substrate binding structures and catalytic mechanism, overview
Mycolicibacterium vanbaalenii
Substrates and Products (Substrate)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
ATP + maltose
-
739666
Mycolicibacterium vanbaalenii
ADP + alpha-maltose-1-phosphate
-
-
-
?
ATP + maltose
-
739666
Mycobacterium tuberculosis
ADP + alpha-maltose-1-phosphate
-
-
-
?
ATP + maltose
-
739666
Mycolicibacterium vanbaalenii DSM 7251
ADP + alpha-maltose-1-phosphate
-
-
-
?
ATP + maltose
-
739666
Mycobacterium tuberculosis ATCC 25618
ADP + alpha-maltose-1-phosphate
-
-
-
?
Subunits
Subunits
Commentary
Organism
More
the N-terminal lobe can be divided into two subdomains: a cap N-terminal subdomain comprising the first 88 amino acid residues and an intermediate subdomain composed of an anti-parallel beta-sheet flanked by two helices. The C-terminal lobe is mostly alpha-helical. While the N-terminal cap subdomain and the C-terminal lobe are predominantly acidic, the intermediate subdomain is enriched in positively charged residues. The N-terminal cap subdomain is composed of three long antiparallel beta-strands forming a curved beta-sheet that encloses the N-terminal alpha-helix and a short two-stranded beta-sheet running perpendicular to the longest beta-sheet axis, on its concave surface. The intermediate subdomain (residues 89-200) contains a central seven-stranded beta-sheet flanked by two alpha-helical segments. A nine-residue linker (residues 201-209) containing a short beta-strand connects the intermediate subdomain and the C-terminal lobe. This last domain is composed of two central 4-helical bundles, a short beta-hairpin and a small two-stranded beta-sheet
Mycolicibacterium vanbaalenii
Temperature Optimum [°C]
Temperature Optimum [°C]
Temperature Optimum Maximum [°C]
Commentary
Organism
30
-
assay at
Mycolicibacterium vanbaalenii
37
-
assay at
Mycobacterium tuberculosis
60
-
-
Mycolicibacterium vanbaalenii
pH Optimum
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
7.5
-
assay at
Mycobacterium tuberculosis
7.5
-
assay at
Mycolicibacterium vanbaalenii
Cofactor
Cofactor
Commentary
Organism
Structure
ATP
-
Mycobacterium tuberculosis
ATP
-
Mycolicibacterium vanbaalenii
Cloned(Commentary) (protein specific)
Commentary
Organism
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)
Mycolicibacterium vanbaalenii
gene mak, phylogenetic analysis, recombinant expression of C-terminally His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
Mycobacterium tuberculosis
Cofactor (protein specific)
Cofactor
Commentary
Organism
Structure
ATP
-
Mycobacterium tuberculosis
ATP
-
Mycolicibacterium vanbaalenii
Crystallization (Commentary) (protein specific)
Crystallization
Organism
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
Mycolicibacterium vanbaalenii
Engineering (protein specific)
Amino acid exchange
Commentary
Organism
D334N
inactive mutant
Mycolicibacterium vanbaalenii
D339N
inactive mutant
Mycobacterium tuberculosis
E324R
site-directed mutagenesis, the mutant shows decreased activity compared to the wild-type enzyme
Mycolicibacterium vanbaalenii
E340R
site-directed mutagenesis, the mutant shows decreased activity compared to the wild-type enzyme
Mycobacterium tuberculosis
K413A
site-directed mutagenesis
Mycolicibacterium vanbaalenii
N137A
site-directed mutagenesis, inactive mutant
Mycolicibacterium vanbaalenii
N145A
site-directed mutagenesis, inactive mutant
Mycobacterium tuberculosis
R334R
site-directed mutagenesis, the mutant shows decreased activity compared to the wild-type enzyme
Mycolicibacterium vanbaalenii
R351A
site-directed mutagenesis, the mutant shows decreased activity compared to the wild-type enzyme
Mycobacterium tuberculosis
S136A
site-directed mutagenesis, the mutant shows highly decreased activity compared to the wild-type enzyme
Mycolicibacterium vanbaalenii
S144A
site-directed mutagenesis, the mutant shows highly decreased activity compared to the wild-type enzyme
Mycobacterium tuberculosis
Y416A
site-directed mutagenesis
Mycolicibacterium vanbaalenii
Y416F
site-directed mutagenesis
Mycolicibacterium vanbaalenii
Y420A
site-directed mutagenesis
Mycolicibacterium vanbaalenii
Y420F
site-directed mutagenesis
Mycolicibacterium vanbaalenii
Metals/Ions (protein specific)
Metals/Ions
Commentary
Organism
Structure
Mg2+
required
Mycobacterium tuberculosis
Mg2+
required, magnesium-binding residue is Asp322
Mycolicibacterium vanbaalenii
Natural Substrates/ Products (Substrates) (protein specific)
Natural Substrates
Organism
Commentary (Nat. Sub.)
Natural Products
Commentary (Nat. Pro.)
Organism (Nat. Pro.)
Reversibility
ATP + maltose
Mycolicibacterium vanbaalenii
-
ADP + alpha-maltose-1-phosphate
-
-
?
ATP + maltose
Mycobacterium tuberculosis
-
ADP + alpha-maltose-1-phosphate
-
-
?
ATP + maltose
Mycolicibacterium vanbaalenii DSM 7251
-
ADP + alpha-maltose-1-phosphate
-
-
?
ATP + maltose
Mycobacterium tuberculosis ATCC 25618
-
ADP + alpha-maltose-1-phosphate
-
-
?
Purification (Commentary) (protein specific)
Commentary
Organism
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
Mycobacterium tuberculosis
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
Mycolicibacterium vanbaalenii
Substrates and Products (Substrate) (protein specific)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
ATP + maltose
-
739666
Mycolicibacterium vanbaalenii
ADP + alpha-maltose-1-phosphate
-
-
-
?
ATP + maltose
-
739666
Mycobacterium tuberculosis
ADP + alpha-maltose-1-phosphate
-
-
-
?
ATP + maltose
-
739666
Mycolicibacterium vanbaalenii DSM 7251
ADP + alpha-maltose-1-phosphate
-
-
-
?
ATP + maltose
-
739666
Mycobacterium tuberculosis ATCC 25618
ADP + alpha-maltose-1-phosphate
-
-
-
?
Subunits (protein specific)
Subunits
Commentary
Organism
More
the N-terminal lobe can be divided into two subdomains: a cap N-terminal subdomain comprising the first 88 amino acid residues and an intermediate subdomain composed of an anti-parallel beta-sheet flanked by two helices. The C-terminal lobe is mostly alpha-helical. While the N-terminal cap subdomain and the C-terminal lobe are predominantly acidic, the intermediate subdomain is enriched in positively charged residues. The N-terminal cap subdomain is composed of three long antiparallel beta-strands forming a curved beta-sheet that encloses the N-terminal alpha-helix and a short two-stranded beta-sheet running perpendicular to the longest beta-sheet axis, on its concave surface. The intermediate subdomain (residues 89-200) contains a central seven-stranded beta-sheet flanked by two alpha-helical segments. A nine-residue linker (residues 201-209) containing a short beta-strand connects the intermediate subdomain and the C-terminal lobe. This last domain is composed of two central 4-helical bundles, a short beta-hairpin and a small two-stranded beta-sheet
Mycolicibacterium vanbaalenii
Temperature Optimum [°C] (protein specific)
Temperature Optimum [°C]
Temperature Optimum Maximum [°C]
Commentary
Organism
30
-
assay at
Mycolicibacterium vanbaalenii
37
-
assay at
Mycobacterium tuberculosis
60
-
-
Mycolicibacterium vanbaalenii
pH Optimum (protein specific)
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
7.5
-
assay at
Mycobacterium tuberculosis
7.5
-
assay at
Mycolicibacterium vanbaalenii
General Information
General Information
Commentary
Organism
evolution
the enzyme belongs to the family of eukaryotic-like kinases (ELKs) with N-terminal domain topologically resembling the cystatin family of protease inhibitors. Phylogenetic analysis, overview
Mycobacterium tuberculosis
evolution
the enzyme belongs to the family of eukaryotic-like kinases (ELKs) with N-terminal domain topologically resembling the cystatin family of protease inhibitors. Phylogenetic analysis, overview
Mycolicibacterium vanbaalenii
metabolism
the enzyme catalyzes the fourth and last step of the GlcE pathway that channels trehalose to glycogen synthesis and is also likely involved in the biosynthesis of two other crucial polymers: intracellular methylglucose lipopolysaccharides and exposed capsular glucan
Mycobacterium tuberculosis
metabolism
the enzyme catalyzes the fourth and last step of the GlcE pathway that channels trehalose to glycogen synthesis and is also likely involved in the biosynthesis of two other crucial polymers: intracellular methylglucose lipopolysaccharides and exposed capsular glucan
Mycolicibacterium vanbaalenii
additional information
the enzyme shows an eukaryotic-like kinase (ELK) fold, similar to methylthioribose kinases and aminoglycoside phosphotransferases, a typical eukaryotic protein kinase-like fold. Subtle structural rearrangements occur upon nucleotide binding in the cleft between the N- and the C-terminal lobes. The enzyme has a phosphate-binding region in the N-terminal lobe that is proposed to act as an anchoring point tethering maltokinase and trehalose isomerase activities to the site of glycogen biosynthesis, ensuring correct regulation of Mak activity and possibly preventing excessive accumulation of maltose 1-phosphate. The enzyme's unusual N-terminal domain, with the 146AMLKV150 motif, containing the conserved phosphate-binding lysine residue, might regulate its phosphotransfer activity and represents the most likely anchoring point for TreS, the upstream enzyme in the pathway. Putative catalytic base is residue Asp305
Mycolicibacterium vanbaalenii
General Information (protein specific)
General Information
Commentary
Organism
evolution
the enzyme belongs to the family of eukaryotic-like kinases (ELKs) with N-terminal domain topologically resembling the cystatin family of protease inhibitors. Phylogenetic analysis, overview
Mycobacterium tuberculosis
evolution
the enzyme belongs to the family of eukaryotic-like kinases (ELKs) with N-terminal domain topologically resembling the cystatin family of protease inhibitors. Phylogenetic analysis, overview
Mycolicibacterium vanbaalenii
metabolism
the enzyme catalyzes the fourth and last step of the GlcE pathway that channels trehalose to glycogen synthesis and is also likely involved in the biosynthesis of two other crucial polymers: intracellular methylglucose lipopolysaccharides and exposed capsular glucan
Mycobacterium tuberculosis
metabolism
the enzyme catalyzes the fourth and last step of the GlcE pathway that channels trehalose to glycogen synthesis and is also likely involved in the biosynthesis of two other crucial polymers: intracellular methylglucose lipopolysaccharides and exposed capsular glucan
Mycolicibacterium vanbaalenii
additional information
the enzyme shows an eukaryotic-like kinase (ELK) fold, similar to methylthioribose kinases and aminoglycoside phosphotransferases, a typical eukaryotic protein kinase-like fold. Subtle structural rearrangements occur upon nucleotide binding in the cleft between the N- and the C-terminal lobes. The enzyme has a phosphate-binding region in the N-terminal lobe that is proposed to act as an anchoring point tethering maltokinase and trehalose isomerase activities to the site of glycogen biosynthesis, ensuring correct regulation of Mak activity and possibly preventing excessive accumulation of maltose 1-phosphate. The enzyme's unusual N-terminal domain, with the 146AMLKV150 motif, containing the conserved phosphate-binding lysine residue, might regulate its phosphotransfer activity and represents the most likely anchoring point for TreS, the upstream enzyme in the pathway. Putative catalytic base is residue Asp305
Mycolicibacterium vanbaalenii
Other publictions for EC 2.7.1.175
No.
1st author
Pub Med
title
organims
journal
volume
pages
year
Activating Compound
Application
Cloned(Commentary)
Crystallization (Commentary)
Engineering
General Stability
Inhibitors
KM Value [mM]
Localization
Metals/Ions
Molecular Weight [Da]
Natural Substrates/ Products (Substrates)
Organic Solvent Stability
Organism
Oxidation Stability
Posttranslational Modification
Purification (Commentary)
Reaction
Renatured (Commentary)
Source Tissue
Specific Activity [micromol/min/mg]
Storage Stability
Substrates and Products (Substrate)
Subunits
Temperature Optimum [°C]
Temperature Range [°C]
Temperature Stability [°C]
Turnover Number [1/s]
pH Optimum
pH Range
pH Stability
Cofactor
Ki Value [mM]
pI Value
IC50 Value
Activating Compound (protein specific)
Application (protein specific)
Cloned(Commentary) (protein specific)
Cofactor (protein specific)
Crystallization (Commentary) (protein specific)
Engineering (protein specific)
General Stability (protein specific)
IC50 Value (protein specific)
Inhibitors (protein specific)
Ki Value [mM] (protein specific)
KM Value [mM] (protein specific)
Localization (protein specific)
Metals/Ions (protein specific)
Molecular Weight [Da] (protein specific)
Natural Substrates/ Products (Substrates) (protein specific)
Organic Solvent Stability (protein specific)
Oxidation Stability (protein specific)
Posttranslational Modification (protein specific)
Purification (Commentary) (protein specific)
Renatured (Commentary) (protein specific)
Source Tissue (protein specific)
Specific Activity [micromol/min/mg] (protein specific)
Storage Stability (protein specific)
Substrates and Products (Substrate) (protein specific)
Subunits (protein specific)
Temperature Optimum [°C] (protein specific)
Temperature Range [°C] (protein specific)
Temperature Stability [°C] (protein specific)
Turnover Number [1/s] (protein specific)
pH Optimum (protein specific)
pH Range (protein specific)
pH Stability (protein specific)
pI Value (protein specific)
Expression
General Information
General Information (protein specific)
Expression (protein specific)
KCat/KM [mM/s]
KCat/KM [mM/s] (protein specific)
739666
Fraga
Structure of mycobacterial mal ...
Mycobacterium tuberculosis, Mycobacterium tuberculosis ATCC 25618, Mycolicibacterium vanbaalenii, Mycolicibacterium vanbaalenii DSM 7251
Sci. Rep.
5
8026
2015
-
-
2
1
15
-
-
-
-
2
-
4
-
6
-
-
2
1
-
-
-
-
4
1
3
-
-
-
2
-
-
2
-
-
-
-
-
2
2
1
15
-
-
-
-
-
-
2
-
4
-
-
-
2
-
-
-
-
4
1
3
-
-
-
2
-
-
-
-
5
5
-
-
-
737320
Roy
Synthesis of alpha-glucan in m ...
Mycobacterium tuberculosis, Mycobacterium tuberculosis ATCC 25618
ACS Chem. Biol.
8
2245-2255
2013
1
-
1
-
3
-
-
1
-
1
3
2
-
7
-
-
1
-
-
-
-
-
2
2
-
-
-
-
1
-
-
1
-
-
-
1
-
1
1
-
3
-
-
-
-
1
-
1
3
2
-
-
-
1
-
-
-
-
2
2
-
-
-
-
1
-
-
-
-
3
3
-
-
-
717432
Mendes
Biochemical characterization o ...
Mycobacterium tuberculosis variant bovis
BMC Biochem.
11
21
2010
-
1
1
-
-
1
1
3
-
5
1
1
-
5
-
-
1
-
-
-
-
1
4
1
1
1
-
-
1
1
-
3
-
-
-
-
1
1
3
-
-
1
-
1
-
3
-
5
1
1
-
-
-
1
-
-
-
1
4
1
1
1
-
-
1
1
-
-
-
2
2
-
-
-
717816
Elbein
Last step in the conversion of ...
Mycolicibacterium smegmatis, Mycolicibacterium smegmatis ATCC 14468
J. Biol. Chem.
285
9803-9812
2010
-
1
-
-
-
-
-
-
-
1
-
-
-
4
-
-
-
-
-
-
-
-
2
-
1
-
-
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1
-
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1
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1
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1
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-
-
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1
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2
-
1
-
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1
-
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-
-
1
1
-
-
-
717739
Jarling
Isolation of mak1 from Actinop ...
Actinoplanes missouriensis, Streptomyces coelicolor, Streptomyces coelicolor A3(2)
J. Basic Microbiol.
44
360-373
2004
-
-
2
-
-
-
-
-
-
1
4
-
-
9
-
-
2
-
-
-
1
-
3
2
1
-
-
-
1
-
-
1
-
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2
1
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-
-
-
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-
1
4
-
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2
-
-
1
-
3
2
1
-
-
-
1
-
-
-
-
1
1
-
-
-
717163
Niehues
Isolation and characterization ...
Actinoplanes missouriensis
Arch. Microbiol.
180
233-239
2003
-
-
-
-
-
-
-
3
-
1
1
-
-
2
-
-
1
-
-
-
-
-
1
1
1
1
-
-
2
-
-
2
-
1
-
-
-
-
2
-
-
-
-
-
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3
-
1
1
-
-
-
-
1
-
-
-
-
1
1
1
1
-
-
2
-
-
1
-
-
-
-
-
-
717600
Drepper
Maltokinase (ATP:maltose 1-pho ...
Actinoplanes sp.
FEBS Lett.
388
177-179
1996
-
-
-
-
-
-
-
-
-
1
-
1
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2
-
-
-
-
-
-
-
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2
-
1
-
-
-
1
-
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1
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-
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-
1
-
-
-
-
-
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-
1
-
1
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-
-
-
-
-
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2
-
1
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1
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