BRENDA - Enzyme Database show

Identification of residues involved in the specificity and regulation of the highly efficient multisubstrate deoxyribonucleoside kinase from Drosophila melanogaster

Knecht, W.; Munch-Petersen, B.; Piskur, J.; J. Mol. Biol. 301, 827-837 (2000)

Data extracted from this reference:

Engineering
EC Number
Amino acid exchange
Commentary
Organism
2.7.1.145
I102M/N117S/M118V/D208N
clones harbouring the mutant enzyme are more sensitive to 3'-azido-2',3'-dideoxythymidine and beta-D-arabinofuranosylcytosine
Drosophila melanogaster
2.7.1.145
I47T/N210D
clones harbouring the mutant enzyme are more sensitive to 3'-azido-2',3'-dideoxythymidine and beta-D-arabinofuranosylcytosine
Drosophila melanogaster
2.7.1.145
M1T/T85A/N121S
clones harbouring the mutant enzyme are more sensitive to 1-beta-D-arabinofuranosylcytosine and 2',3'-dideoxycytidine
Drosophila melanogaster
2.7.1.145
N210D/L239P
clones harbouring the mutant enzyme are more sensitive to 3'-azido-2',3'-dideoxythymidine and beta-D-arabinofuranosylcytosine
Drosophila melanogaster
2.7.1.145
N38D/N64D
clones harbouring the mutant enzyme are more sensitive specifically to 3'-azido-2',3'-dideoxythymidine
Drosophila melanogaster
2.7.1.145
N45D/N64D
clones harbouring the mutant enzyme are more sensitive to 3'-azido-2',3'-dideoxythymidine and other nucleoside analogs; mutant enzyme with the largest und universal increase in sensitivity of all mutants tested. 316fold increase to 3'-azido-2',3'-dideoxythymidine, more than 11fold to 2',3'-dideoxycytidine, and 3.2fold increase in sensitivity to beta-D-arabinofuranosylcytosine and 2',3'-dideoxyadenosine. Higher Km-values for native substrates than wild-type enzyme and Vmax-values are substantially lower, decrease in feedback inhibition by TTP, Km and Vmax values for 3'-azido-2',3'-dideoxythymidine and Km value for 2',3'-dideoxycytidine are nearly unchanged
Drosophila melanogaster
2.7.1.145
N59D/M118V/Y179H
clones harbouring the mutant enzyme are more sensitive to 3'-azido-2',3'-dideoxythymidine and beta-D-arabinofuranosylcytosine
Drosophila melanogaster
2.7.1.145
N64S/L68S/M69L
clones harbouring the mutant enzyme are more sensitive specifically to 3'-azido-2',3'-dideoxythymidine
Drosophila melanogaster
2.7.1.145
T12A/V84A/N213S
clones harbouring the mutant enzyme are more sensitive to 1-beta-D-arabinofuranosylcytosine
Drosophila melanogaster
2.7.1.145
T85A
clones harbouring the mutant enzyme are more sensitive to 1-beta-D-arabinofuranosylcytosine and 2',3'-dideoxycytidine
Drosophila melanogaster
2.7.1.145
V84A
clones harbouring the mutant enzyme are more sensitive to 1-beta-D-arabinofuranosylcytosine
Drosophila melanogaster
Inhibitors
EC Number
Inhibitors
Commentary
Organism
Structure
2.7.1.145
TTP
mutant enzyme N45D/N64D shows a decrease in feedback inhibition compared to the wild-type enzyme
Drosophila melanogaster
KM Value [mM]
EC Number
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
2.7.1.145
0.0072
-
3'-azido-2',3'-dideoxythymidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
0.0083
-
3'-azido-2',3'-dideoxythymidine
-
Drosophila melanogaster
2.7.1.145
0.0242
-
thymidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
0.0964
-
2'-deoxycytidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
1.093
-
2',3'-dideoxycytidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
1.124
-
2',3'-dideoxycytidine
-
Drosophila melanogaster
2.7.1.145
2.004
-
2'-deoxyguanosine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
3.16
-
2'-deoxyadenosine
mutant enzyme N45D/N64D
Drosophila melanogaster
Organism
EC Number
Organism
Primary Accession No. (UniProt)
Commentary
Textmining
2.7.1.145
Drosophila melanogaster
-
-
-
Substrates and Products (Substrate)
EC Number
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
2.7.1.145
(E)-5-(2-bromovinyl)-2'-deoxyuridine + ATP
54% of the activity with thymidine
636844
Drosophila melanogaster
(E)-5-(2-bromovinyl)-2'-deoxyuridine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
1-beta-D-arabinofuranosylcytosine + ATP
58.2% of the activity with thymidine
636844
Drosophila melanogaster
1-beta-D-arabinofuranosylcytosine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
1-beta-D-arabinofuranosylthymine + ATP
53.9% of the activity with thymidine
636844
Drosophila melanogaster
1-beta-D-arabinofuranosylthymine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
2',3'-dideoxyadenosine + ATP
0.9% of the activity with thymidine
636844
Drosophila melanogaster
2',3'-dideoxyadenosine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
2',3'-dideoxycytidine + ATP
-
636844
Drosophila melanogaster
2',3'-dideoxycytidine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
2'-deoxyadenosine + ATP
-
636844
Drosophila melanogaster
2'-deoxyadenosine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
2'-deoxycytidine + ATP
-
636844
Drosophila melanogaster
2'-deoxycytidine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
2'-deoxyguanosine + ATP
-
636844
Drosophila melanogaster
2'-deoxyguanosine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
2'-deoxyuridine + ATP
113% of the activity with thymidine
636844
Drosophila melanogaster
2'-deoxyuridine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
2-chloro-2'-deoxyadenosine + ATP
126% of the activity with thymidine
636844
Drosophila melanogaster
2-chloro-2'-deoxyadenosine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
3'-azido-2',3'-dideoxythymidine + ATP
-
636844
Drosophila melanogaster
3'-azido-2',3'-dideoxythymidine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
5-fluoro-2'-deoxyuridine + ATP
92% of the activity with thymidine
636844
Drosophila melanogaster
5-fluoro-2'-deoxyuridine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
adenosine + ATP
0.82% of the activity with thymidine
636844
Drosophila melanogaster
adenosine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
cytidine + ATP
16% of the activity with thymidine
636844
Drosophila melanogaster
cytidine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
guanosine + ATP
1.18% of the activity with thymidine
636844
Drosophila melanogaster
guanosine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
thymidine + ATP
-
636844
Drosophila melanogaster
thymidine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
uridine + ATP
6.45% of the activity with thymidine
636844
Drosophila melanogaster
uridine 5'-phosphate + ADP
-
-
-
?
Turnover Number [1/s]
EC Number
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
2.7.1.145
0.036
-
3'-azido-2',3'-dideoxythymidine
-
Drosophila melanogaster
2.7.1.145
0.052
-
3'-azido-2',3'-dideoxythymidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
0.076
-
2'-deoxyguanosine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
0.54
-
2',3'-dideoxycytidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
0.828
-
2'-deoxyadenosine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
1.22
-
thymidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
4.03
-
2'-deoxycytidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
4.2
-
2',3'-dideoxycytidine
-
Drosophila melanogaster
2.7.1.145
14.2
-
thymidine
pH 7.8, 22°C, recombinant enzyme
Drosophila melanogaster
2.7.1.145
15.1
-
2'-deoxyguanosine
pH 7.8, 22°C, recombinant enzyme
Drosophila melanogaster
2.7.1.145
16.5
-
2'-deoxycytidine
pH 7.8, 22°C, recombinant enzyme
Drosophila melanogaster
2.7.1.145
20.6
-
2'-deoxyadenosine
pH 7.8, 22°C, recombinant enzyme
Drosophila melanogaster
Engineering (protein specific)
EC Number
Amino acid exchange
Commentary
Organism
2.7.1.145
I102M/N117S/M118V/D208N
clones harbouring the mutant enzyme are more sensitive to 3'-azido-2',3'-dideoxythymidine and beta-D-arabinofuranosylcytosine
Drosophila melanogaster
2.7.1.145
I47T/N210D
clones harbouring the mutant enzyme are more sensitive to 3'-azido-2',3'-dideoxythymidine and beta-D-arabinofuranosylcytosine
Drosophila melanogaster
2.7.1.145
M1T/T85A/N121S
clones harbouring the mutant enzyme are more sensitive to 1-beta-D-arabinofuranosylcytosine and 2',3'-dideoxycytidine
Drosophila melanogaster
2.7.1.145
N210D/L239P
clones harbouring the mutant enzyme are more sensitive to 3'-azido-2',3'-dideoxythymidine and beta-D-arabinofuranosylcytosine
Drosophila melanogaster
2.7.1.145
N38D/N64D
clones harbouring the mutant enzyme are more sensitive specifically to 3'-azido-2',3'-dideoxythymidine
Drosophila melanogaster
2.7.1.145
N45D/N64D
clones harbouring the mutant enzyme are more sensitive to 3'-azido-2',3'-dideoxythymidine and other nucleoside analogs; mutant enzyme with the largest und universal increase in sensitivity of all mutants tested. 316fold increase to 3'-azido-2',3'-dideoxythymidine, more than 11fold to 2',3'-dideoxycytidine, and 3.2fold increase in sensitivity to beta-D-arabinofuranosylcytosine and 2',3'-dideoxyadenosine. Higher Km-values for native substrates than wild-type enzyme and Vmax-values are substantially lower, decrease in feedback inhibition by TTP, Km and Vmax values for 3'-azido-2',3'-dideoxythymidine and Km value for 2',3'-dideoxycytidine are nearly unchanged
Drosophila melanogaster
2.7.1.145
N59D/M118V/Y179H
clones harbouring the mutant enzyme are more sensitive to 3'-azido-2',3'-dideoxythymidine and beta-D-arabinofuranosylcytosine
Drosophila melanogaster
2.7.1.145
N64S/L68S/M69L
clones harbouring the mutant enzyme are more sensitive specifically to 3'-azido-2',3'-dideoxythymidine
Drosophila melanogaster
2.7.1.145
T12A/V84A/N213S
clones harbouring the mutant enzyme are more sensitive to 1-beta-D-arabinofuranosylcytosine
Drosophila melanogaster
2.7.1.145
T85A
clones harbouring the mutant enzyme are more sensitive to 1-beta-D-arabinofuranosylcytosine and 2',3'-dideoxycytidine
Drosophila melanogaster
2.7.1.145
V84A
clones harbouring the mutant enzyme are more sensitive to 1-beta-D-arabinofuranosylcytosine
Drosophila melanogaster
Inhibitors (protein specific)
EC Number
Inhibitors
Commentary
Organism
Structure
2.7.1.145
TTP
mutant enzyme N45D/N64D shows a decrease in feedback inhibition compared to the wild-type enzyme
Drosophila melanogaster
KM Value [mM] (protein specific)
EC Number
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
2.7.1.145
0.0072
-
3'-azido-2',3'-dideoxythymidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
0.0083
-
3'-azido-2',3'-dideoxythymidine
-
Drosophila melanogaster
2.7.1.145
0.0242
-
thymidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
0.0964
-
2'-deoxycytidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
1.093
-
2',3'-dideoxycytidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
1.124
-
2',3'-dideoxycytidine
-
Drosophila melanogaster
2.7.1.145
2.004
-
2'-deoxyguanosine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
3.16
-
2'-deoxyadenosine
mutant enzyme N45D/N64D
Drosophila melanogaster
Substrates and Products (Substrate) (protein specific)
EC Number
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
2.7.1.145
(E)-5-(2-bromovinyl)-2'-deoxyuridine + ATP
54% of the activity with thymidine
636844
Drosophila melanogaster
(E)-5-(2-bromovinyl)-2'-deoxyuridine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
1-beta-D-arabinofuranosylcytosine + ATP
58.2% of the activity with thymidine
636844
Drosophila melanogaster
1-beta-D-arabinofuranosylcytosine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
1-beta-D-arabinofuranosylthymine + ATP
53.9% of the activity with thymidine
636844
Drosophila melanogaster
1-beta-D-arabinofuranosylthymine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
2',3'-dideoxyadenosine + ATP
0.9% of the activity with thymidine
636844
Drosophila melanogaster
2',3'-dideoxyadenosine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
2',3'-dideoxycytidine + ATP
-
636844
Drosophila melanogaster
2',3'-dideoxycytidine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
2'-deoxyadenosine + ATP
-
636844
Drosophila melanogaster
2'-deoxyadenosine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
2'-deoxycytidine + ATP
-
636844
Drosophila melanogaster
2'-deoxycytidine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
2'-deoxyguanosine + ATP
-
636844
Drosophila melanogaster
2'-deoxyguanosine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
2'-deoxyuridine + ATP
113% of the activity with thymidine
636844
Drosophila melanogaster
2'-deoxyuridine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
2-chloro-2'-deoxyadenosine + ATP
126% of the activity with thymidine
636844
Drosophila melanogaster
2-chloro-2'-deoxyadenosine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
3'-azido-2',3'-dideoxythymidine + ATP
-
636844
Drosophila melanogaster
3'-azido-2',3'-dideoxythymidine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
5-fluoro-2'-deoxyuridine + ATP
92% of the activity with thymidine
636844
Drosophila melanogaster
5-fluoro-2'-deoxyuridine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
adenosine + ATP
0.82% of the activity with thymidine
636844
Drosophila melanogaster
adenosine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
cytidine + ATP
16% of the activity with thymidine
636844
Drosophila melanogaster
cytidine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
guanosine + ATP
1.18% of the activity with thymidine
636844
Drosophila melanogaster
guanosine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
thymidine + ATP
-
636844
Drosophila melanogaster
thymidine 5'-phosphate + ADP
-
-
-
?
2.7.1.145
uridine + ATP
6.45% of the activity with thymidine
636844
Drosophila melanogaster
uridine 5'-phosphate + ADP
-
-
-
?
Turnover Number [1/s] (protein specific)
EC Number
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
2.7.1.145
0.036
-
3'-azido-2',3'-dideoxythymidine
-
Drosophila melanogaster
2.7.1.145
0.052
-
3'-azido-2',3'-dideoxythymidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
0.076
-
2'-deoxyguanosine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
0.54
-
2',3'-dideoxycytidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
0.828
-
2'-deoxyadenosine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
1.22
-
thymidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
4.03
-
2'-deoxycytidine
mutant enzyme N45D/N64D
Drosophila melanogaster
2.7.1.145
4.2
-
2',3'-dideoxycytidine
-
Drosophila melanogaster
2.7.1.145
14.2
-
thymidine
pH 7.8, 22°C, recombinant enzyme
Drosophila melanogaster
2.7.1.145
15.1
-
2'-deoxyguanosine
pH 7.8, 22°C, recombinant enzyme
Drosophila melanogaster
2.7.1.145
16.5
-
2'-deoxycytidine
pH 7.8, 22°C, recombinant enzyme
Drosophila melanogaster
2.7.1.145
20.6
-
2'-deoxyadenosine
pH 7.8, 22°C, recombinant enzyme
Drosophila melanogaster