BRENDA - Enzyme Database show
show all sequences of 2.7.1.145

Structural basis for the changed substrate specificity of Drosophila melanogaster deoxyribonucleoside kinase mutant N64D

Welin, M.; Skovgaard, T.; Knecht, W.; Zhu, C.; Berenstein, D.; Munch-Petersen, B.; Piskur, J.; Eklund, H.; FEBS J. 272, 3733-3742 (2005)

Data extracted from this reference:

Cloned(Commentary)
Commentary
Organism
expression of the wild-type and mutant enzymes in enzyme-deficient Escherichia coli strain KY895
Drosophila melanogaster
Crystallization (Commentary)
Crystallization
Organism
purified recombinant mutant N64D in complex with dT or dTTP, counter diffusion and vapour diffusion methods, 20 mg/ml enzyme with 10 mM dT, is mixed with cyrstallization solution containing 0.15 M MES, pH 6.5, 0.3 M lithium sulfate, and 27,5% w/v PEG 2000 monomethylether, or 10 mg/ml enzyme with 5 mM dTTP is mixed with 0.1 M MES, pH 6.5, 0.16 M lithium sulfate, and 25% v/v PEG 2000 monomethylether, 2 weeks, 15°C, X-ray diffraction structure determination and analysis at 3.1 A and 2.2 A resolution, respectively
Drosophila melanogaster
Engineering
Amino acid exchange
Commentary
Organism
additional information
expression of the enzyme in enzyme-deficient Escherichia coli strain KY895 confers resistnce against 3'-azido-2',3'-didehydro-3'-deoxythymidine at more than 0.1 mM
Drosophila melanogaster
N45D
site-directed mutagenesis, decreased activity with natural substrates
Drosophila melanogaster
N45D/N64D
decreased activity with natural substrates, decreased feedback inhibition by dTTP compared to the wild-type enzyme
Drosophila melanogaster
N64D
site-directed mutagenesis, highly decreased activity with natural substrates, decreased feedback inhibition by dTTP compared to the wild-type enzyme
Drosophila melanogaster
Inhibitors
Inhibitors
Commentary
Organism
Structure
dTTP
competitive versus ATP, feedback inhibition, binding at the active site, binding structure of mutant N64D, modelling
Drosophila melanogaster
KM Value [mM]
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.0009
-
2'-deoxycytidine
recombinant mutant N45D
Drosophila melanogaster
0.0009
-
2'-deoxythymidine
recombinant mutants N45D
Drosophila melanogaster
0.0111
-
3'-azido-2',3'-didehydro-3'-deoxythymidine
recombinant mutant N64D
Drosophila melanogaster
0.0117
-
3'-azido-2',3'-didehydro-3'-deoxythymidine
recombinant mutant N45D
Drosophila melanogaster
0.0232
-
2'-deoxythymidine
recombinant mutant N64D
Drosophila melanogaster
0.118
-
2'-deoxycytidine
recombinant mutant N64D
Drosophila melanogaster
0.119
-
2'-deoxyadenosine
recombinant mutant N45D
Drosophila melanogaster
0.412
-
2'-deoxyguanosine
recombinant mutant N45D
Drosophila melanogaster
3.82
-
2'-deoxyadenosine
recombinant mutant N64D
Drosophila melanogaster
20.35
-
2'-deoxyguanosine
recombinant mutant N64D
Drosophila melanogaster
Metals/Ions
Metals/Ions
Commentary
Organism
Structure
Mg2+
-
Drosophila melanogaster
Natural Substrates/ Products (Substrates)
Natural Substrates
Organism
Commentary (Nat. Sub.)
Natural Products
Commentary (Nat. Pro.)
Organism (Nat. Pro.)
Reversibility
ATP + 2'-deoxyadenosine
Drosophila melanogaster
-
ADP + 2'-deoxyadenosine 5'-phosphate
-
-
?
ATP + 2'-deoxycytidine
Drosophila melanogaster
-
ADP + 2'-deoxycytidine 5'-phosphate
-
-
?
ATP + 2'-deoxyguanosine
Drosophila melanogaster
-
ADP + 2'-deoxyguanosine 5'-phosphate
-
-
?
ATP + 2'-deoxythymidine
Drosophila melanogaster
-
ADP + 2'-deoxythymidine 5'-phosphate
-
-
?
Organism
Organism
Primary Accession No. (UniProt)
Commentary
Textmining
Drosophila melanogaster
-
-
-
Specific Activity [micromol/min/mg]
Specific Activity Minimum [µmol/min/mg]
Specific Activity Maximum [µmol/min/mg]
Commentary
Organism
additional information
-
-
Drosophila melanogaster
Substrates and Products (Substrate)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
ATP + 2'-deoxyadenosine
-
661702
Drosophila melanogaster
ADP + 2'-deoxyadenosine 5'-phosphate
-
-
-
?
ATP + 2'-deoxycytidine
-
661702
Drosophila melanogaster
ADP + 2'-deoxycytidine 5'-phosphate
-
-
-
?
ATP + 2'-deoxyguanosine
-
661702
Drosophila melanogaster
ADP + 2'-deoxyguanosine 5'-phosphate
-
-
-
?
ATP + 2'-deoxythymidine
-
661702
Drosophila melanogaster
ADP + 2'-deoxythymidine 5'-phosphate
-
-
-
?
ATP + 2'-deoxythymidine
substrate binding structure of mutant N64D
661702
Drosophila melanogaster
ADP + 2'-deoxythymidine 5'-phosphate
-
-
-
?
ATP + 3'-azido-2',3'-didehydro-3'-deoxythymidine
-
661702
Drosophila melanogaster
ADP + 3'-azido-2',3'-didehydro-3'-deoxythymidine 5'-phosphate
-
-
-
?
ATP + aciclovir
-
661702
Drosophila melanogaster
ADP + aciclovir phosphate
-
-
-
?
Turnover Number [1/s]
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
0.03
-
3'-azido-2',3'-didehydro-3'-deoxythymidine
recombinant mutant N45D
Drosophila melanogaster
0.037
-
3'-azido-2',3'-didehydro-3'-deoxythymidine
recombinant mutant N64D
Drosophila melanogaster
0.12
-
2'-deoxyguanosine
recombinant mutant N64D
Drosophila melanogaster
0.4
-
2'-deoxyadenosine
recombinant mutant N64D
Drosophila melanogaster
0.82
-
2'-deoxythymidine
recombinant mutant N64D
Drosophila melanogaster
1.3
-
2'-deoxyguanosine
recombinant mutant N45D
Drosophila melanogaster
1.6
-
2'-deoxycytidine
recombinant mutants N45D and N64D
Drosophila melanogaster
1.83
-
2'-deoxyadenosine
recombinant mutant N45D
Drosophila melanogaster
2.6
-
2'-deoxythymidine
recombinant mutant N45D
Drosophila melanogaster
Cofactor
Cofactor
Commentary
Organism
Structure
ATP
-
Drosophila melanogaster
Ki Value [mM]
Ki Value [mM]
Ki Value maximum [mM]
Inhibitor
Commentary
Organism
Structure
0.0047
-
dTTP
uncompetitive versus 2'-deoxythymidine, recombinant wild-type enzyme
Drosophila melanogaster
0.0163
-
dTTP
competitive versus ATP, recombinant wild-type enzyme
Drosophila melanogaster
0.52
-
dTTP
uncompetitive versus 2'-deoxythymidine, recombinant mutant N64D
Drosophila melanogaster
0.829
-
dTTP
competitive versus ATP, recombinant mutant N64D
Drosophila melanogaster
Cloned(Commentary) (protein specific)
Commentary
Organism
expression of the wild-type and mutant enzymes in enzyme-deficient Escherichia coli strain KY895
Drosophila melanogaster
Cofactor (protein specific)
Cofactor
Commentary
Organism
Structure
ATP
-
Drosophila melanogaster
Crystallization (Commentary) (protein specific)
Crystallization
Organism
purified recombinant mutant N64D in complex with dT or dTTP, counter diffusion and vapour diffusion methods, 20 mg/ml enzyme with 10 mM dT, is mixed with cyrstallization solution containing 0.15 M MES, pH 6.5, 0.3 M lithium sulfate, and 27,5% w/v PEG 2000 monomethylether, or 10 mg/ml enzyme with 5 mM dTTP is mixed with 0.1 M MES, pH 6.5, 0.16 M lithium sulfate, and 25% v/v PEG 2000 monomethylether, 2 weeks, 15°C, X-ray diffraction structure determination and analysis at 3.1 A and 2.2 A resolution, respectively
Drosophila melanogaster
Engineering (protein specific)
Amino acid exchange
Commentary
Organism
additional information
expression of the enzyme in enzyme-deficient Escherichia coli strain KY895 confers resistnce against 3'-azido-2',3'-didehydro-3'-deoxythymidine at more than 0.1 mM
Drosophila melanogaster
N45D
site-directed mutagenesis, decreased activity with natural substrates
Drosophila melanogaster
N45D/N64D
decreased activity with natural substrates, decreased feedback inhibition by dTTP compared to the wild-type enzyme
Drosophila melanogaster
N64D
site-directed mutagenesis, highly decreased activity with natural substrates, decreased feedback inhibition by dTTP compared to the wild-type enzyme
Drosophila melanogaster
Inhibitors (protein specific)
Inhibitors
Commentary
Organism
Structure
dTTP
competitive versus ATP, feedback inhibition, binding at the active site, binding structure of mutant N64D, modelling
Drosophila melanogaster
Ki Value [mM] (protein specific)
Ki Value [mM]
Ki Value maximum [mM]
Inhibitor
Commentary
Organism
Structure
0.0047
-
dTTP
uncompetitive versus 2'-deoxythymidine, recombinant wild-type enzyme
Drosophila melanogaster
0.0163
-
dTTP
competitive versus ATP, recombinant wild-type enzyme
Drosophila melanogaster
0.52
-
dTTP
uncompetitive versus 2'-deoxythymidine, recombinant mutant N64D
Drosophila melanogaster
0.829
-
dTTP
competitive versus ATP, recombinant mutant N64D
Drosophila melanogaster
KM Value [mM] (protein specific)
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.0009
-
2'-deoxycytidine
recombinant mutant N45D
Drosophila melanogaster
0.0009
-
2'-deoxythymidine
recombinant mutants N45D
Drosophila melanogaster
0.0111
-
3'-azido-2',3'-didehydro-3'-deoxythymidine
recombinant mutant N64D
Drosophila melanogaster
0.0117
-
3'-azido-2',3'-didehydro-3'-deoxythymidine
recombinant mutant N45D
Drosophila melanogaster
0.0232
-
2'-deoxythymidine
recombinant mutant N64D
Drosophila melanogaster
0.118
-
2'-deoxycytidine
recombinant mutant N64D
Drosophila melanogaster
0.119
-
2'-deoxyadenosine
recombinant mutant N45D
Drosophila melanogaster
0.412
-
2'-deoxyguanosine
recombinant mutant N45D
Drosophila melanogaster
3.82
-
2'-deoxyadenosine
recombinant mutant N64D
Drosophila melanogaster
20.35
-
2'-deoxyguanosine
recombinant mutant N64D
Drosophila melanogaster
Metals/Ions (protein specific)
Metals/Ions
Commentary
Organism
Structure
Mg2+
-
Drosophila melanogaster
Natural Substrates/ Products (Substrates) (protein specific)
Natural Substrates
Organism
Commentary (Nat. Sub.)
Natural Products
Commentary (Nat. Pro.)
Organism (Nat. Pro.)
Reversibility
ATP + 2'-deoxyadenosine
Drosophila melanogaster
-
ADP + 2'-deoxyadenosine 5'-phosphate
-
-
?
ATP + 2'-deoxycytidine
Drosophila melanogaster
-
ADP + 2'-deoxycytidine 5'-phosphate
-
-
?
ATP + 2'-deoxyguanosine
Drosophila melanogaster
-
ADP + 2'-deoxyguanosine 5'-phosphate
-
-
?
ATP + 2'-deoxythymidine
Drosophila melanogaster
-
ADP + 2'-deoxythymidine 5'-phosphate
-
-
?
Specific Activity [micromol/min/mg] (protein specific)
Specific Activity Minimum [µmol/min/mg]
Specific Activity Maximum [µmol/min/mg]
Commentary
Organism
additional information
-
-
Drosophila melanogaster
Substrates and Products (Substrate) (protein specific)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
ATP + 2'-deoxyadenosine
-
661702
Drosophila melanogaster
ADP + 2'-deoxyadenosine 5'-phosphate
-
-
-
?
ATP + 2'-deoxycytidine
-
661702
Drosophila melanogaster
ADP + 2'-deoxycytidine 5'-phosphate
-
-
-
?
ATP + 2'-deoxyguanosine
-
661702
Drosophila melanogaster
ADP + 2'-deoxyguanosine 5'-phosphate
-
-
-
?
ATP + 2'-deoxythymidine
-
661702
Drosophila melanogaster
ADP + 2'-deoxythymidine 5'-phosphate
-
-
-
?
ATP + 2'-deoxythymidine
substrate binding structure of mutant N64D
661702
Drosophila melanogaster
ADP + 2'-deoxythymidine 5'-phosphate
-
-
-
?
ATP + 3'-azido-2',3'-didehydro-3'-deoxythymidine
-
661702
Drosophila melanogaster
ADP + 3'-azido-2',3'-didehydro-3'-deoxythymidine 5'-phosphate
-
-
-
?
ATP + aciclovir
-
661702
Drosophila melanogaster
ADP + aciclovir phosphate
-
-
-
?
Turnover Number [1/s] (protein specific)
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
0.03
-
3'-azido-2',3'-didehydro-3'-deoxythymidine
recombinant mutant N45D
Drosophila melanogaster
0.037
-
3'-azido-2',3'-didehydro-3'-deoxythymidine
recombinant mutant N64D
Drosophila melanogaster
0.12
-
2'-deoxyguanosine
recombinant mutant N64D
Drosophila melanogaster
0.4
-
2'-deoxyadenosine
recombinant mutant N64D
Drosophila melanogaster
0.82
-
2'-deoxythymidine
recombinant mutant N64D
Drosophila melanogaster
1.3
-
2'-deoxyguanosine
recombinant mutant N45D
Drosophila melanogaster
1.6
-
2'-deoxycytidine
recombinant mutants N45D and N64D
Drosophila melanogaster
1.83
-
2'-deoxyadenosine
recombinant mutant N45D
Drosophila melanogaster
2.6
-
2'-deoxythymidine
recombinant mutant N45D
Drosophila melanogaster
Other publictions for EC 2.7.1.145
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)
738739
Kamiya
In vivo selection of active de ...
Drosophila melanogaster
J. Biotechnol.
228
52-57
2016
-
-
1
-
-
-
1
-
-
1
-
1
-
3
-
-
1
-
-
-
-
-
3
-
1
-
-
-
1
-
-
1
-
-
-
-
-
1
1
-
-
-
-
1
-
-
-
1
-
1
-
-
-
1
-
-
-
-
3
-
1
-
-
-
1
-
-
-
-
1
1
-
-
-
738781
Slot Christiansen
Non-viral deoxyribonucleoside ...
Anopheles gambiae, Bombyx mori, Drosophila melanogaster
J. Genet. Genomics
42
235-248
2015
-
2
-
1
-
-
-
-
1
3
-
18
-
3
-
-
-
-
-
-
-
-
33
-
-
-
-
-
-
-
-
3
-
-
-
-
2
-
3
1
-
-
-
-
-
-
1
3
-
18
-
-
-
-
-
-
-
-
33
-
-
-
-
-
-
-
-
-
-
7
7
-
-
-
739211
Jiang
Tanshinone IIA enhances bystan ...
Drosophila melanogaster
Oncol. Rep.
34
1487-1493
2015
-
1
1
-
-
-
-
-
4
-
-
1
-
3
-
-
-
-
-
1
-
-
3
-
-
-
-
-
-
-
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1
-
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1
1
1
-
-
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-
-
-
-
4
-
-
1
-
-
-
-
-
1
-
-
3
-
-
-
-
-
-
-
-
-
-
1
1
-
-
-
737404
Serra
-
Immobilized Drosophila melanog ...
Drosophila melanogaster
Adv. Synth. Catal.
356
563-570
2014
-
1
-
-
1
-
-
-
-
1
-
1
-
1
-
-
-
-
-
-
-
-
14
-
1
-
-
-
1
-
-
1
-
-
-
-
1
-
1
-
1
-
-
-
-
-
-
1
-
1
-
-
-
-
-
-
-
-
14
-
1
-
-
-
1
-
-
-
-
-
-
-
-
-
703396
Betham
Use of Drosophila deoxynucleos ...
Drosophila melanogaster
DNA Repair
9
153-160
2010
-
-
1
-
-
-
-
-
-
-
-
-
-
4
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
1
-
-
-
-
-
1
1
-
-
-
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-
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-
1
-
-
-
-
-
-
-
-
-
-
1
1
-
-
-
721775
Ito
Cytotoxic effect of Drosophila ...
Drosophila melanogaster
Biol. Pharm. Bull.
33
1223-1227
2010
-
-
1
-
-
-
-
-
-
-
-
-
-
5
-
-
-
-
-
-
-
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1
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-
-
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-
-
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-
702047
Knecht
Drosophila melanogaster deoxyr ...
Drosophila melanogaster
Biochem. Biophys. Res. Commun.
382
430-433
2009
-
-
-
1
-
-
-
1
-
-
-
-
-
3
-
-
-
-
-
-
-
-
1
-
-
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-
1
-
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1
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1
1
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1
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-
-
1
-
-
-
-
1
-
-
-
-
-
-
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-
1
1
692289
Mikkelsen
Structural studies of nucleosi ...
Drosophila melanogaster
FEBS J.
275
2151-2160
2008
-
-
-
1
-
-
-
-
-
-
-
-
-
2
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
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1
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-
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-
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-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
694476
Clausen
A multisubstrate deoxyribonucl ...
Arabidopsis thaliana
Nucleic Acids Symp. Ser.
52
489-490
2008
-
-
1
-
-
-
-
-
-
-
-
-
-
2
-
-
-
-
-
-
-
-
7
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
7
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
673584
Egeblad-Welin
Functional studies of active-s ...
Drosophila melanogaster
FEBS J.
274
1542-1551
2007
-
-
-
1
7
-
-
18
-
-
-
-
-
2
-
-
-
-
-
-
-
-
5
-
-
-
-
18
-
-
-
-
-
-
-
-
-
-
-
1
7
-
-
-
-
18
-
-
-
-
-
-
-
-
-
-
-
-
5
-
-
-
-
18
-
-
-
-
-
-
-
-
-
-
673848
Knecht
Drosophila deoxyribonucleoside ...
Drosophila melanogaster
Gene Ther.
14
1278-1286
2007
-
1
1
-
2
-
-
-
-
-
-
-
-
4
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
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-
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-
-
-
-
1
1
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-
2
-
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-
-
-
-
-
-
-
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1
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Mitochondrial expression of th ...
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672971
Legent
In vivo analysis of Drosophila ...
Drosophila melanogaster
Cell Cycle
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676247
Kamiya
Transient expression of Drosop ...
Drosophila melanogaster
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Welin
Structural basis for the chang ...
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661731
Piskur
Animal deoxyribonucleoside kin ...
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662990
Solaroli
Investigation of the substrate ...
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660855
Krawiec
Inorganic tripolyphosphate (PP ...
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2003
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661072
Mikkelsen
Structural basis for feedback ...
Drosophila melanogaster
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Solaroli
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9
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Knecht
Mosquito has a single multisub ...
Anopheles gambiae
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662985
Krawiec
Ability of adenosine-2'(3')-de ...
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2003
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661453
Eriksson
Structure and function of cell ...
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661583
Knecht
A few amino acid substitutions ...
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Knecht
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Zheng
Nucleoside analog cytotoxicity ...
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Johansson
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636842
Munch-Petersen
Functional expression of a mul ...
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1
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636843
Balzarini
The multifunctional deoxynucle ...
Drosophila melanogaster
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Knecht
Identification of residues inv ...
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636841
Johansson
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636839
Munch-Petersen
The single deoxynucleoside kin ...
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636840
Munch-Petersen
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