Cloned (Comment) | Organism |
---|---|
expressed in Escherichia coli BL21(DE3) cells | Human gammaherpesvirus 4 |
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 | 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 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 | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | required, Mg2+ plays a role mainly in substrate binding | Human gammaherpesvirus 4 |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Human gammaherpesvirus 4 | P03195 | - |
- |
Purification (Comment) | Organism |
---|---|
Ni-NTA column chromatography | Human gammaherpesvirus 4 |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
dUTP + H2O | - |
Human gammaherpesvirus 4 | dUMP + diphosphate | - |
? |
Subunits | Comment | Organism |
---|---|---|
monomer | - |
Human gammaherpesvirus 4 |
Synonyms | Comment | Organism |
---|---|---|
deoxyuridine 5'-triphosphate | - |
Human gammaherpesvirus 4 |
dUTPase | - |
Human gammaherpesvirus 4 |
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 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 |