Information on EC 2.7.6.2 - thiamine diphosphokinase

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

EC NUMBER
COMMENTARY hide
2.7.6.2
-
RECOMMENDED NAME
GeneOntology No.
thiamine diphosphokinase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + thiamine = AMP + thiamine diphosphate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
diphosphate transfer
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Metabolic pathways
-
-
thiamine diphosphate biosynthesis III (Staphylococcus)
-
-
thiamine diphosphate biosynthesis IV (eukaryotes)
-
-
Thiamine metabolism
-
-
thiamine salvage III
-
-
thiamine salvage IV (yeast)
-
-
vitamin B1 metabolism
-
-
SYSTEMATIC NAME
IUBMB Comments
ATP:thiamine diphosphotransferase
-
CAS REGISTRY NUMBER
COMMENTARY hide
9026-24-8
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
rainbow trout
UniProt
Manually annotated by BRENDA team
strain 12442
-
-
Manually annotated by BRENDA team
strain 12442
-
-
Manually annotated by BRENDA team
serovar typhimurium LT2
-
-
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
ATP + thiamine
AMP + thiamine diphosphate
show the reaction diagram
CTP + thiamine
CMP + thiamine diphosphate
show the reaction diagram
GTP + thiamine
GMP + thiamine diphosphate
show the reaction diagram
oxythiamine + ATP
oxythiamine diphosphate + AMP
show the reaction diagram
-
-
-
-
?
pyrithiamine + ATP
pyrithiamine diphosphate + AMP
show the reaction diagram
thiamin + ATP
thiamindiphosphate + AMP
show the reaction diagram
-
-
-
?
thiamine + ATP
thiamine diphosphate + AMP
show the reaction diagram
UTP + thiamine
UMP + thiamine diphosphate
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
ATP + thiamine
AMP + thiamine diphosphate
show the reaction diagram
thiamin + ATP
thiamindiphosphate + AMP
show the reaction diagram
F4IV16
-
-
-
?
thiamine + ATP
thiamine diphosphate + AMP
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
myo-Inositol 1-diphosphate
-
coenzyme
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ca2+
-
divalent cation required, 7.3% of the activation with Mg2+
additional information
-
ligand binding structures, overview
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
ALT-711
-
low-affinity inhibitor, mixed-mode inhibition with a major role for competitive inhibition
AMP
-
0.5 mM, 8% inhibition
Butylthiamine
-
competitive with respect to thiamine
Chloroethylthiamine
-
weak
diphosphate
ethylthiamine
-
competitive with respect to thiamine
iodoacetamide
KCN
-
little effect
molybdate
-
little effect
NaF
-
little effect
Oxythiamine
pyrithiamine
thiamine diphosphate
Thiamine monophosphate
-
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
myo-inositol
-
stimulates
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0041 - 18.3
ATP
7.6
CTP
-
pH 8.1, 37C
0.021 - 2.6
GTP
4.5
MgATP2-
-
-
0.062
pyrithiamine
-
-
0.00015 - 2.9
thiamine
11.2
UTP
-
pH 8.1, 37C
additional information
additional information
-
-
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0002 - 0.094
ATP
0.045
thiamin
Homo sapiens
-
pH 7.5, 37C, mutant enzyme Q96E
0.0002 - 0.094
thiamine
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2.8
diphosphate
-
-
0.019
pyrithiamine
-
-
0.2
Thiamine monophosphate
-
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.000061
-
-
0.000367
-
recombinant enzyme
0.00165
-
-
0.009
-
PfTPK3, with 4 mM magnesium ions in the enzyme assay
0.017
-
PfTPK2, with 4 mM magnesium ions in the enzyme assay; substrate: pyrithiamine, PfTPK
0.027
-
PfTPK, with 4 mM magnesium ions in the enzyme assay
0.0602
-
-
999
-
no activity with thiochrome
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
isozymes TPK1 or TPK2
7.1 - 7.3
-
-
8.3 - 9.3
-
in 0.028 M phosphate/glycylglycine buffer
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.4 - 8
-
pH 6.4: about 40% of maximal activity, pH 8.0: about 50% of maximal activity
6.8 - 9.3
-
pH 6.8: about 65% of maximal activity, pH 8.3-9.3: optimum
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
45
AtTPK1; isozyme TPK1
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37 - 50
37C: optimal activity of TPK2, 50C: TPK2 is completely inactive
50
AtTPK2 is completely inactive
55
AtTPK1 is still active
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4.2
-
isoelectric focusing
4.5
-
isoelectric focusing
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
-
low expression
Manually annotated by BRENDA team
-
highly expressed
Manually annotated by BRENDA team
-
low expression
Manually annotated by BRENDA team
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
-
Peroxisomes are devoid of thiamine pyrophosphokinase activity.
Manually annotated by BRENDA team
additional information
PDB
SCOP
CATH
ORGANISM
UNIPROT
Bacillus subtilis (strain 168)
Bacteroides thetaiotaomicron (strain ATCC 29148 / DSM 2079 / NCTC 10582 / E50 / VPI-5482)
Candida albicans (strain SC5314 / ATCC MYA-2876)
Candida albicans (strain SC5314 / ATCC MYA-2876)
Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30000
-
gel filtration
38000
-
for PfTPK3, SDS-PAGE analysis followed by Western blotting
43000
-
for PfTPK2, SDS-PAGE analysis followed by Western blotting
44000
-
gel filtration
47000
-
for PfTPK, SDS-PAGE analysis followed by Western blotting
49000
-
gel filtration
61000
-
gel filtration
96000
-
aggregate of the functional dimer, equilibrium sedimentation
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
x * 28000-30000, SDS-PAGE
homodimer
-
-
homotetramer
-
x-ray crystallography
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified recombinant enzyme in complex with ATP analogue AMP-PNP and thiamine, hanging drop vapor diffusion method, mixing of 500 nl of protein solution with 500 nl of reservoir solution containing 17.5-20% PEG 4000, 0.2 M MgCl2 , 0.1 M Tris, and 20% glycerol, pH 7.0-pH 7.5, and equilibration against 1 ml or reservoir solution, X-ray diffraction structure determination and analysis at 1.96-2.1 A resolution
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complexed with thiamin diphosphate-Mg2+ or thiamin-Mg2+, hanging drop vapor diffusion method, using 1.15-1.3 M sodium citrate, 0.1 M Na-HEPES, pH 8.6, at 22C
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Crystallization experiments were set up by incubating mouse TPK with pyrithiamine and MgATP2- prior to adding the equilibrium mixture of enzyme, substrate, and products to the previously determined crystallization conditions.
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recombinant enzyme, hanging drop vapor diffusion method
-
hanging-drop vapor diffusion method, crystals of the complex of native recombinant enzyme with thiamine
-
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50
inactivation of isozyme TPK2
55
isozyme TPK1 shows remaining activity
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
the enzyme becomes increasingly unstable with progressive purification
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
nickel-chelate chromatography
-
partial
recombinant CA1462 from Escherichia coli
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Escherichia coli strain KL21 (DE3)pLysS is transformed with the expression vector coding for a histidine-tagged wild-type or mutant hTPK1
-
expressed in Escherichia coli BL21 cells
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gene CA1462, expression in Escherichia coli
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isozymes TPK1 and TPK2, DNA and amino acid sequence determination and analysis, phylogenetic analysis, expression in Escherichia coli strain BL21, expression of fluorescent tagged isozymes in cytosol of mesophyll protoplasts
overexpression in Escherichia coli
-
TPK1 gene, located in 7q34-36, genotyping in 964 individuals from 220 families of European ancestry revealing an association between maternal rs228584 genotype and offspring birth weight
-
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
during embryogenesis, total thiamin diphosphokinase transcripts increase abruptly in early development, and decrease to about half of the peak shortly after hatching
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D100N
-
mutation reduces turnover-number markedly
D133N
-
mutation causes a selective decrease in the ratio of turnover-number to Km-value for thiamine
D71N
-
mutation reduces turnover-number markedly
D73N
-
mutation reduces turnover-number markedly
Q96E
-
mutation causes an 2.5fold increase in the ratio of turnover-number to Km-value for thiamine compared to the wild-type
R131G
-
mutation decreases the ratio of turnover-number to Km-value for ATP
S74A
-
mutation causes a 1.4fold increase in turnover number, the Km-value for ATP is 2fold that of the wild-type enzyme
T99A
-
mutation decreases the ratio of turnover-number to Km-value for ATP
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
medicine
-
Malaria is a devastating and quite often deadly parasitic disease, resulting in significant public health problems in the tropics. The inevitable emergence of drug-resistant forms requires the continuous discovery and development of new antimalarials. These drugs should only target the parasite, with only harmless or no effects on the human host. Thus, ideal drug targets would be peculiarities in the parasite metabolism, such as vitamin B1 and B6 biosyntheses that are absent in its host.
additional information
-
Administration of oxythiamine to animals produces primarily metabolic aberrations such as lethargy and anorexia while the administration of pyrithiamine to animals produces neurological problems similar to Wernicke-Korsakoff Syndrome.
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