Cloned (Comment) | Organism |
---|---|
expressed in Escherichia coli BL21(DE3) cells | Nitrosomonas europaea |
Crystallization (Comment) | Organism |
---|---|
hanging drop vapor diffusion method, selenomethionine-substituted protein, using 3.5 M sodium formate as a precipitation agent and 0.1 M Bis-Tris propane buffer, pH 7.0, at 20°C | Nitrosomonas europaea |
Protein Variants | Comment | Organism |
---|---|---|
K52R | the His-tagged mutant loses 90-99% of its activity (catalytic efficiency is at least 1000times lower) compared to the wild type enzyme. Mn2+ does not induce a significant activation of this mutant | Nitrosomonas europaea |
K85A | the His-tagged mutant is about 10times less active than the wild type enzyme, but the optimal conditions for activity are essentially the same. The mutant is more strongly activated by Mn2+ than by Mg2+, and the inhibitory effects of Ca2+ and Zn2+ are less pronounced | Nitrosomonas europaea |
K8A | the His-tagged mutant remains highly active with tripolyphosphate as substrate and Mg2+ as activator, with a catalytic efficiency close to that of the recombinant wild type enzyme | Nitrosomonas europaea |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
Cd2+ | - |
Nitrosomonas europaea | |
Cu2+ | - |
Nitrosomonas europaea | |
Zn2+ | complete inhibition at 0.005 mM | Nitrosomonas europaea |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.021 | - |
tripolyphosphate | His-tagged wild type enzyme, at pH 9.7 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
0.04 | - |
tripolyphosphate | His-tagged wild type enzyme, at pH 9.7 and 50°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
0.058 | - |
tripolyphosphate | His-tagged wild type enzyme, at pH 7.1 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
0.1 | - |
tripolyphosphate | untagged wild type enzyme, at pH 9.7 and 50°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
0.191 | - |
tripolyphosphate | His-tagged mutant enzyme K52R, at pH 9.7 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
0.39 | - |
tripolyphosphate | His-tagged mutant enzyme K8A, at pH 9.7 and 50°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
0.72 | - |
tripolyphosphate | His-tagged mutant enzyme K85A, at pH 9.7 and 50°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
0.8 | - |
ATP | His-tagged wild type enzyme, at pH 8.1 and 50°C, in the presence of 10 mM Mn2+ | Nitrosomonas europaea | |
1.2 | - |
ATP | His-tagged mutant enzyme K8A, at pH 10.1 and 50°C, in the presence of 10 mM Mn2+ | Nitrosomonas europaea | |
2.6 | - |
tripolyphosphate | His-tagged mutant enzyme K85A, at pH 9.7 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
74 | - |
tripolyphosphate | His-tagged mutant enzyme K8A, at pH 9.7 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Co2+ | Co2+ is a better activator than Mn2+, although the maximum activity is less than 10% that measured in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
Mg2+ | the enzyme hydrolyzes tripolyphosphate with high catalytic efficiency in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
Mn2+ | very poor substituent for Mg2+ in the pH range 7.0-10, but there is a significant Mn2+-dependent activity at pH 10.0-10.5. Half-maximum activation is obtained at 0.4 mM | Nitrosomonas europaea |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
19000 | - |
2 * 19000, SDS-PAGE | Nitrosomonas europaea |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Nitrosomonas europaea | - |
- |
- |
Purification (Comment) | Organism |
---|---|
His-Trap column chromatography, gel filtration | Nitrosomonas europaea |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
ATP + H2O | low affinity | Nitrosomonas europaea | ? | - |
? | |
additional information | the enzyme has low affinity for CTP, ATP or thiamine triphosphate. Nucleoside triphosphatase activity is negligible in the presence of 5 mM Mg2+, but a small activity is observed in the presence of 1 mM Mn2+, in particular with GTP. Guanosine 5-tetraphosphate and long chain polyphosphate (containing about 65 phosphate residues) are not hydrolyzed. The enzyme has no adenylyl cyclase activity | Nitrosomonas europaea | ? | - |
? | |
tripolyphosphate + H2O | very good substrate with Mg2+ as activator. The enzyme has a strong preference for linear tripolyphosphate compared with cyclic trimetaphosphate and to the linear tetraphosphate | Nitrosomonas europaea | diphosphate + phosphate | - |
? |
Subunits | Comment | Organism |
---|---|---|
homodimer | 2 * 19000, SDS-PAGE | Nitrosomonas europaea |
Synonyms | Comment | Organism |
---|---|---|
inorganic triphosphatase | - |
Nitrosomonas europaea |
triphosphate tunnel metalloenzyme | - |
Nitrosomonas europaea |
TTM | - |
Nitrosomonas europaea |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
50 | 55 | - |
Nitrosomonas europaea |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.28 | - |
tripolyphosphate | His-tagged mutant enzyme K8A, at pH 9.7 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
0.36 | - |
ATP | His-tagged wild type enzyme, at pH 8.1 and 50°C, in the presence of 10 mM Mn2+ | Nitrosomonas europaea | |
0.98 | - |
tripolyphosphate | His-tagged mutant enzyme K52R, at pH 9.7 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
3.96 | - |
ATP | His-tagged mutant enzyme K8A, at pH 10.1 and 50°C, in the presence of 10 mM Mn2+ | Nitrosomonas europaea | |
10 | - |
tripolyphosphate | His-tagged mutant enzyme K85A, at pH 9.7 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
21 | - |
tripolyphosphate | His-tagged mutant enzyme K85A, at pH 9.7 and 50°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
60 | - |
tripolyphosphate | His-tagged wild type enzyme, at pH 7.1 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
76 | - |
tripolyphosphate | His-tagged wild type enzyme, at pH 9.7 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
288 | - |
tripolyphosphate | His-tagged wild type enzyme, at pH 9.7 and 50°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
887 | - |
tripolyphosphate | His-tagged mutant enzyme K8A, at pH 9.7 and 50°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
7900 | - |
tripolyphosphate | untagged wild type enzyme, at pH 9.7 and 50°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
9.7 | - |
around pH 9.7 | Nitrosomonas europaea |
IC50 Value | IC50 Value Maximum | Comment | Organism | Inhibitor | Structure |
---|---|---|---|---|---|
0.0015 | - |
His-tagged wild type enzyme, at pH 9.7 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | Zn2+ | |
0.01 | - |
His-tagged wild type enzyme, at pH 9.7 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | Cd2+ | |
0.02 | - |
His-tagged wild type enzyme, at pH 9.7 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | Cu2+ |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.45 | - |
ATP | His-tagged wild type enzyme, at pH 8.1 and 50°C, in the presence of 10 mM Mn2+ | Nitrosomonas europaea | |
3.3 | - |
ATP | His-tagged mutant enzyme K8A, at pH 10.1 and 50°C, in the presence of 10 mM Mn2+ | Nitrosomonas europaea | |
4 | - |
tripolyphosphate | His-tagged mutant enzyme K85A, at pH 9.7 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
5.1 | - |
tripolyphosphate | His-tagged mutant enzyme K52R, at pH 9.7 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
29 | - |
tripolyphosphate | His-tagged mutant enzyme K85A, at pH 9.7 and 50°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
190 | - |
tripolyphosphate | His-tagged mutant enzyme K8A, at pH 9.7 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
2300 | - |
tripolyphosphate | His-tagged mutant enzyme K8A, at pH 9.7 and 50°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
3300 | - |
tripolyphosphate | His-tagged wild type enzyme, at pH 7.1 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
3600 | - |
tripolyphosphate | His-tagged wild type enzyme, at pH 9.7 and 37°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
7200 | - |
tripolyphosphate | His-tagged wild type enzyme, at pH 9.7 and 50°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea | |
79000 | - |
tripolyphosphate | untagged wild type enzyme, at pH 9.7 and 50°C, in the presence of 5 mM Mg2+ | Nitrosomonas europaea |