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Literature summary extracted from

  • Mikoulinskaia, G.; Taran, S.; Skoblov, Y.; Feofanov, S.
    The study of the bacteriophage T5 deoxynucleoside monophosphate kinase active site by site-directed mutagenesis (2013), Russ. J. Bioorg. Chem., 39, 607-618.
No PubMed abstract available

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.7.4.13 gene dnk, recombinant expression of wild-type and mutant enzymes in Escherichia coli strains BL21(DE3) and XL10Gold Tequintavirus T5

Protein Variants

EC Number Protein Variants Comment Organism
2.7.4.13 D16N site-directed mutagenesis of the P loop-related residue, the mutant shows reduced activity and altered substrate specificity compared to the wild-type enzyme Tequintavirus T5
2.7.4.13 D170N site-directed mutagenesis of the core residue, almost inactive mutant Tequintavirus T5
2.7.4.13 E176Q site-directed mutagenesis of the core residue, almost inactive mutant Tequintavirus T5
2.7.4.13 G137A site-directed mutagenesis of the dNMP-binding residue, the mutant shows highly reduced activity and altered substrate specificity compared to the wild-type enzyme Tequintavirus T5
2.7.4.13 K131E site-directed mutagenesis, of the dNMP-binding residue, the mutant shows slightly increased activity and altered substrate specificity compared to the wild-type enzyme Tequintavirus T5
2.7.4.13 additional information all enzyme mutants but T17S show an increased pH optimum compared to the wild-type enzyme Tequintavirus T5
2.7.4.13 Q134A site-directed mutagenesis of the dNMP-binding residue, the mutant shows reduced activity and altered substrate specificity compared to the wild-type enzyme Tequintavirus T5
2.7.4.13 R130K site-directed mutagenesis of the dNMP-binding residue, almost inactive mutant Tequintavirus T5
2.7.4.13 R172I site-directed mutagenesis of the core residue, almost inactive mutant Tequintavirus T5
2.7.4.13 S13A site-directed mutagenesis of the P loop-related residue, the mutant shows reduced activity and altered substrate specificity compared to the wild-type enzyme Tequintavirus T5
2.7.4.13 T138A site-directed mutagenesis of the dNMP-binding residue, the mutant shows reduced activity and altered substrate specificity compared to the wild-type enzyme Tequintavirus T5
2.7.4.13 T17N site-directed mutagenesis of the P loop-related residue, the mutant shows highly reduced activity and altered substrate specificity compared to the wild-type enzyme Tequintavirus T5
2.7.4.13 T17S site-directed mutagenesis, of the P loop-related residue, the mutant shows reduced activity compared to the wild-type enzyme Tequintavirus T5
2.7.4.13 W150A site-directed mutagenesis of the core residue Tequintavirus T5
2.7.4.13 W150F site-directed mutagenesis of the conserved residue, the mutant shows unaltered activity compared to the wild-type enzyme Tequintavirus T5

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
2.7.4.13 Mg2+ required Tequintavirus T5

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.7.4.13 ATP + dAMP Tequintavirus T5
-
ADP + dADP
-
?
2.7.4.13 ATP + dCMP Tequintavirus T5
-
ADP + dCDP
-
?
2.7.4.13 ATP + dGMP Tequintavirus T5
-
ADP + dGDP
-
?
2.7.4.13 ATP + dTMP Tequintavirus T5
-
ADP + dTDP
-
?

Organism

EC Number Organism UniProt Comment Textmining
2.7.4.13 Tequintavirus T5 Q6QGP4 gene dnk
-

Purification (Commentary)

EC Number Purification (Comment) Organism
2.7.4.13 recombinant wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by anion exchange and affinity chromatography Tequintavirus T5

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
2.7.4.13 additional information
-
specific activties of wild-type enzyme and enzyme mutants with different dNMP acceptor substrates, overview Tequintavirus T5
2.7.4.13 25.78
-
pH 7.5, 37°C, recombinant wild-type enzyme, substrate dCMP Tequintavirus T5
2.7.4.13 36.54
-
pH 7.5, 37°C, recombinant wild-type enzyme, substrate dGMP Tequintavirus T5
2.7.4.13 41.24
-
pH 7.5, 37°C, recombinant wild-type enzyme, substrate dTMP Tequintavirus T5
2.7.4.13 73.17
-
pH 7.5, 37°C, recombinant wild-type enzyme, substrate dAMP Tequintavirus T5

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.7.4.13 ATP + dAMP
-
Tequintavirus T5 ADP + dADP
-
?
2.7.4.13 ATP + dAMP best substrate dAMP, wild-type enzyme Tequintavirus T5 ADP + dADP
-
?
2.7.4.13 ATP + dCMP
-
Tequintavirus T5 ADP + dCDP
-
?
2.7.4.13 ATP + dCMP 35.2% of the activity with dAMP, wild-type enzyme Tequintavirus T5 ADP + dCDP
-
?
2.7.4.13 ATP + dGMP
-
Tequintavirus T5 ADP + dGDP
-
?
2.7.4.13 ATP + dGMP 49.9% of the activity with dAMP, wild-type enzyme Tequintavirus T5 ADP + dGDP
-
?
2.7.4.13 ATP + dTMP
-
Tequintavirus T5 ADP + dTDP
-
?
2.7.4.13 ATP + dTMP 56.4% of the activity with dAMP, wild-type enzyme Tequintavirus T5 ADP + dTDP
-
?
2.7.4.13 additional information substrate specificities of wild-type and mutant enzymes, structure-function analysis, overview Tequintavirus T5 ?
-
?

Subunits

EC Number Subunits Comment Organism
2.7.4.13 monomer
-
Tequintavirus T5

Synonyms

EC Number Synonyms Comment Organism
2.7.4.13 deoxyribonucleoside monophosphate kinase
-
Tequintavirus T5
2.7.4.13 dNMPK
-
Tequintavirus T5
2.7.4.13 T5 bacteriophage dNMPK
-
Tequintavirus T5

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
2.7.4.13 25
-
assay at Tequintavirus T5

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
2.7.4.13 7
-
wild-type enzyme and mutant T17S Tequintavirus T5
2.7.4.13 7.5 8.5 all enzyme mutants but T17S Tequintavirus T5

Cofactor

EC Number Cofactor Comment Organism Structure
2.7.4.13 ATP
-
Tequintavirus T5

General Information

EC Number General Information Comment Organism
2.7.4.13 additional information essential catalytic site residue, active site modelling, overview. The charged amino acid residues of the NMP binding domain and the presence of an OH-group at position 17 are important for the catalytic activity. Arginine residues at positions 130 and 172 are involved in the binding to the donor gamma-phosphoryl and acceptor alpha-phosphoryl groups, as well as the aspartic acid residue at position 16 of the ATP-binding site (P-loop), in the binding to some acceptors, first of all dTMP. The NMPK-P loop, or a glycine enriched loop, forms an anion hole interacting with donor phosphoryl groups. It contains a GX1X2X3X4GKX5T(S) consensus called the Walker A motif Tequintavirus T5