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Literature summary for 3.1.3.32 extracted from
- Distinct enzymic functional groups are required for the phosphomonoesterase and phosphodiesterase activities of Clostridium thermocellum polynucleotide kinase/phosphatase (2006), J. Biol. Chem., 281, 19251-19259.
Cloned(Commentary)
| Cloned (Comment) | Organism |
|---|---|
| expressed in Escherichia coli as His-tagged fusion protein | Acetivibrio thermocellus |
Protein Variants
| Protein Variants | Comment | Organism |
|---|---|---|
| D236A | decreased activity compared to the wild type enzyme | Acetivibrio thermocellus |
| D236E | decreased activity compared to the wild type enzyme | Acetivibrio thermocellus |
| D236N | similar activity compared to the wild type enzyme | Acetivibrio thermocellus |
| D392E | decreased activity compared to the wild type enzyme | Acetivibrio thermocellus |
| D392N | decreased activity compared to the wild type enzyme | Acetivibrio thermocellus |
| H189A | decreased activity compared to the wild type enzyme | Acetivibrio thermocellus |
| H189D | decreased activity compared to the wild type enzyme | Acetivibrio thermocellus |
| H189E | decreased activity compared to the wild type enzyme | Acetivibrio thermocellus |
| H189Q | decreased activity compared to the wild type enzyme | Acetivibrio thermocellus |
| H264A | decreased activity compared to the wild type enzyme | Acetivibrio thermocellus |
| H264N | increased activity compared to the wild type enzyme | Acetivibrio thermocellus |
| H264Q | decreased activity compared to the wild type enzyme | Acetivibrio thermocellus |
| H376D | decreased activity compared to the wild type enzyme | Acetivibrio thermocellus |
| H376N | decreased activity compared to the wild type enzyme | Acetivibrio thermocellus |
| R237A | decreased activity compared to the wild type enzyme | Acetivibrio thermocellus |
| R237K | increased activity compared to the wild type enzyme | Acetivibrio thermocellus |
| R237Q | decreased activity compared to the wild type enzyme | Acetivibrio thermocellus |
KM Value [mM]
| KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|
| 1 | - |
p-nitrophenyl phosphate | mutant enzyme H376N, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus |  |
| 1.3 | - |
p-nitrophenyl phosphate | mutant enzyme D392E, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 1.6 | - |
bis-p-nitrophenyl phosphate | mutant enzyme D236A, at 45°C, in 50 mM Tris-HCl (pH 8.0) 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 2.2 | - |
bis-p-nitrophenyl phosphate | mutant enzyme D236N, at 45°C, in 50 mM Tris-HCl (pH 8.0) 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 3.4 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H264A, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 3.6 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H189D, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 4.9 | - |
p-nitrophenyl phosphate | mutant enzyme D236A, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 5 | - |
p-nitrophenyl phosphate | mutant enzyme D236N, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 5 | - |
bis-p-nitrophenyl phosphate | mutant enzyme D236N, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 5 | - |
bis-p-nitrophenyl phosphate | mutant enzyme D392E, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 5 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H376D, at 45°C, in 50 mM Tris-HCl (pH 8.0) 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 6.3 | - |
bis-p-nitrophenyl phosphate | wild type enzyme, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 6.6 | - |
p-nitrophenyl phosphate | mutant enzyme D236E, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 6.6 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H189A, at 45°C, in 50 mM Tris-HCl (pH 8.0) 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 7.2 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H376N, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 7.4 | - |
bis-p-nitrophenyl phosphate | mutant enzyme D392N, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 9.6 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H264N, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 11 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H264Q, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 12 | - |
bis-p-nitrophenyl phosphate | mutant enzyme D236A, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 12 | - |
bis-p-nitrophenyl phosphate | mutant enzyme R237K, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 12 | - |
p-nitrophenyl phosphate | wild type enzyme, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 13 | - |
p-nitrophenyl phosphate | mutant enzyme R237K, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 18 | - |
bis-p-nitrophenyl phosphate | mutant enzyme R237A, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 18 | - |
bis-p-nitrophenyl phosphate | mutant enzyme R237Q, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 24 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H264A, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 26 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H189Q, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 38 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H264Q, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 58 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H189E, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 75 | - |
p-nitrophenyl phosphate | mutant enzyme H264N, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 88 | - |
bis-p-nitrophenyl phosphate | wild type enzyme, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
Metals/Ions
| Metals/Ions | Comment | Organism | Structure |
|---|---|---|---|
| Mn2+ | required for hydrolysis of bis-p-nitrophenyl phosphate | Acetivibrio thermocellus | |
| Ni2+ | required for hydrolysis of bis-p-nitrophenyl phosphate | Acetivibrio thermocellus | |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Acetivibrio thermocellus | - |
- |
- |
Purification (Commentary)
| Purification (Comment) | Organism |
|---|---|
| nickel-agarose chromatography | Acetivibrio thermocellus |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| bis-p-nitrophenyl phosphate + H2O | - |
Acetivibrio thermocellus | p-nitrophenyl phosphate + phosphate | - |
? | |
| p-nitrophenyl phosphate + H2O | - |
Acetivibrio thermocellus | p-nitrophenol + phosphate | - |
? | |
Synonyms
| Synonyms | Comment | Organism |
|---|---|---|
| PNKP | belongs to the dinuclear metallophosphoesterase superfamily | Acetivibrio thermocellus |
| polynucleotide kinase/phosphatase | multifunctional enzyme that catalyzes the phosphorylation of 5'-OH termini of DNA or RNA polynucleotides and the dephosphorylation of 3'-phosphate and 2'-phosphate ribonucleotides | Acetivibrio thermocellus |
Turnover Number [1/s]
| Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|
| 0.26 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H264Q, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 0.52 | - |
bis-p-nitrophenyl phosphate | mutant enzyme D392N, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 0.92 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H264Q, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 1.1 | - |
bis-p-nitrophenyl phosphate | mutant enzyme R237A, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 1.23 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H264A, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 1.7 | - |
bis-p-nitrophenyl phosphate | wild type enzyme, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 1.88 | - |
p-nitrophenyl phosphate | mutant enzyme D236E, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 2.13 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H376D, at 45°C, in 50 mM Tris-HCl (pH 8.0) 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 2.5 | - |
p-nitrophenyl phosphate | mutant enzyme R237K, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 2.53 | - |
bis-p-nitrophenyl phosphate | mutant enzyme D236A, at 45°C, in 50 mM Tris-HCl (pH 8.0) 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 3 | 6 | bis-p-nitrophenyl phosphate | mutant enzyme H189D, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 3.13 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H376N, at 45°C, in 50 mM Tris-HCl (pH 8.0),0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 3.33 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H189Q, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 3.42 | - |
p-nitrophenyl phosphate | mutant enzyme D392E, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 3.63 | - |
bis-p-nitrophenyl phosphate | mutant enzyme R237K, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 3.72 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H189A, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 4.05 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H376N, at 45°C, in 50 mM Tris-HCl (pH 8.0),0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 5.25 | - |
p-nitrophenyl phosphate | mutant enzyme H376N, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 8.8 | - |
bis-p-nitrophenyl phosphate | mutant enzyme D392E, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 9.33 | - |
bis-p-nitrophenyl phosphate | mutant enzyme D236N, at 45°C, in 50 mM Tris-HCl (pH 8.0) 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 9.58 | - |
p-nitrophenyl phosphate | mutant enzyme D236A, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 12 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H264N, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 13.77 | - |
bis-p-nitrophenyl phosphate | mutant enzyme D392N, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 15 | - |
p-nitrophenyl phosphate | wild type enzyme, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 16.9 | - |
bis-p-nitrophenyl phosphate | wild type enzyme, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 18 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H264A, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 19.33 | - |
bis-p-nitrophenyl phosphate | mutant enzyme R237A, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 23.17 | - |
bis-p-nitrophenyl phosphate | wild type enzyme, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 30.83 | - |
p-nitrophenyl phosphate | wild type enzyme, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 33 | - |
bis-p-nitrophenyl phosphate | mutant enzyme R237Q, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM NiCl2 | Acetivibrio thermocellus | |
| 38.33 | - |
p-nitrophenyl phosphate | mutant enzyme D236N, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 43.5 | - |
bis-p-nitrophenyl phosphate | mutant enzyme D236A, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 52.33 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H264Q, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 65.83 | - |
bis-p-nitrophenyl phosphate | mutant enzyme D236N, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 117.5 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H189E, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 146.7 | - |
p-nitrophenyl phosphate | mutant enzyme H264N, at 45°C, in 50 mM Tris-HCl (pH 7.5), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
| 253.3 | - |
bis-p-nitrophenyl phosphate | mutant enzyme H264A, at 45°C, in 50 mM Tris-HCl (pH 8.0), 0.5 mM MnCl2 | Acetivibrio thermocellus | |
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