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Literature summary for 3.6.1.23 extracted from

  • Freeman, L.; Buisson, M.; Tarbouriech, N.; Van der Heyden, A.; Labbe, P.; Burmeister, W.P.
    The flexible motif V of Epstein-Barr virus deoxyuridine 5'-triphosphate pyrophosphatase is essential for catalysis (2009), J. Biol. Chem., 284, 25280-25289.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli BL21(DE3) cells Human gammaherpesvirus 4

Crystallization (Commentary)

Crystallization (Comment) Organism
vapor diffusion method, using 0.3 M ammonium sulfate, 25% (w/v) polyethylene glycol 3350, 0.1 M HEPES, pH 7.0 (for mutant enzymes D131S/G78D and D131N), or using 150 mM malic acid, pH 7.5, and 25% (w/v) PEG 3350 (for mutant enzyme C4S), or using 0.1 M Tris, pH 8.5, 20% (w/v) polyethylene glycol 3350, 200 mM Li2SO4 (for mutant enzyme DELTAV, a 257-residue construct with the last 22 C-terminal residues removed) Human gammaherpesvirus 4

Protein Variants

Protein Variants Comment Organism
C4S the mutant shows increased activity compared to the wild type enzyme Human gammaherpesvirus 4
D131E the mutant is completely devoid of enzymatic activity Human gammaherpesvirus 4
D131N the mutant shows decreased activity compared to the wild type enzyme Human gammaherpesvirus 4
D131S the mutant shows decreased activity compared to the wild type enzyme Human gammaherpesvirus 4
D131S/G78D the mutant shows decreased activity compared to the wild type enzyme Human gammaherpesvirus 4
F273A the deletion of the flexible C-terminal tail carrying motif V results in a protein completely devoid of enzymatic activity Human gammaherpesvirus 4
R268A the mutant shows decreased activity compared to the wild type enzyme Human gammaherpesvirus 4

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.0006
-
dUTP mutant enzyme C4S, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
0.00071
-
dUTP wild type enzyme, without MgCl2, with dithiothreitol, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
0.0008
-
dUTP wild type enzyme, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
0.0052
-
dUTP mutant enzyme D131S, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
0.006
-
dUTP wild type enzyme, without MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
0.007
-
dUTP mutant enzyme D131N, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
0.008
-
dUTP mutant enzyme R268A, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
0.009
-
dUTP mutant enzyme D131S/G78D, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required, Mg2+ plays a role mainly in substrate binding Human gammaherpesvirus 4

Organism

Organism UniProt Comment Textmining
Human gammaherpesvirus 4 P03195
-
-

Purification (Commentary)

Purification (Comment) Organism
Ni-NTA column chromatography Human gammaherpesvirus 4

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
dUTP + H2O
-
Human gammaherpesvirus 4 dUMP + diphosphate
-
?

Subunits

Subunits Comment Organism
monomer
-
Human gammaherpesvirus 4

Synonyms

Synonyms Comment Organism
deoxyuridine 5'-triphosphate
-
Human gammaherpesvirus 4
dUTPase
-
Human gammaherpesvirus 4

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.04
-
dUTP mutant enzyme D131N, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
0.04
-
dUTP mutant enzyme D131S, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
0.06
-
dUTP mutant enzyme R268A, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
0.18
-
dUTP mutant enzyme D131S/G78D, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
1.2
-
dUTP wild type enzyme, without MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
1.4
-
dUTP wild type enzyme, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
1.6
-
dUTP wild type enzyme, without MgCl2, with dithiothreitol, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
1.7
-
dUTP mutant enzyme C4S, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
6
-
dUTP mutant enzyme D131N, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
8
-
dUTP mutant enzyme D131S, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
8
-
dUTP mutant enzyme R268A, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
20
-
dUTP mutant enzyme D131S/G78D, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
200
-
dUTP wild type enzyme, without MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
1800
-
dUTP wild type enzyme, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
2300
-
dUTP wild type enzyme, without MgCl2, with dithiothreitol, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4
2800
-
dUTP mutant enzyme C4S, with 1 mM MgCl2, in 0.5 mM bicine-NaOH, pH 7.8, at 25°C Human gammaherpesvirus 4