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Literature summary for 3.1.4.1 extracted from
- A new member of the alkaline phosphatase superfamily with a formylglycine nucleophile: structural and kinetic characterisation of a phosphonate monoester hydrolase/phosphodiesterase from Rhizobium leguminosarum (2008), J. Mol. Biol., 384, 120-136.
Cloned(Commentary)
| Cloned (Comment) | Organism |
|---|---|
| expressed in Escherichia coli Tuner(DE3) cells | Rhizobium leguminosarum |
Protein Variants
| Protein Variants | Comment | Organism |
|---|---|---|
| C57A | mutant shows severely reduced activity towards p-nitrophenyl phenylphosphonate compared to the wild type enzyme | Rhizobium leguminosarum |
| C57S | mutant shows reduced activity towards p-nitrophenyl phenylphosphonate compared to the wild type enzyme | Rhizobium leguminosarum |
| H218A | mutant shows reduced activity towards p-nitrophenyl phenylphosphonate compared to the wild type enzyme | Rhizobium leguminosarum |
| K337A | mutant shows reduced activity towards p-nitrophenyl phenylphosphonate compared to the wild type enzyme | Rhizobium leguminosarum |
| N78A | mutant shows reduced activity towards p-nitrophenyl phenylphosphonate compared to the wild type enzyme | Rhizobium leguminosarum |
| Q13A | mutant shows increased activity towards p-nitrophenyl phenylphosphonate compared to the wild type enzyme | Rhizobium leguminosarum |
| T107A | mutant shows reduced activity towards p-nitrophenyl phenylphosphonate compared to the wild type enzyme | Rhizobium leguminosarum |
| Y105A | mutant shows reduced activity towards p-nitrophenyl phenylphosphonate compared to the wild type enzyme | Rhizobium leguminosarum |
KM Value [mM]
| KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|
| 0.31 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme T107A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum |  |
| 1.7 | - |
p-nitrophenyl ethyl phosphate | mutant enzyme C57S, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 2.3 | - |
p-nitrophenyl ethyl phosphate | wild type enzyme, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 2.8 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme C57A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 2.9 | - |
p-nitrophenyl ethyl phosphate | mutant enzyme C57A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 3 | - |
p-Nitrophenyl phenylphosphonate | wild type enzyme, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 3.8 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme C57S, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 4 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme N78A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 4.8 | - |
p-nitrophenyl ethyl phosphate | mutant enzyme T107A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 14 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme K337A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 15 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme H218A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 28 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme Y105A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 43 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme Q13A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 45 | - |
p-nitrophenyl ethyl phosphate | mutant enzyme N78A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 50 | - |
p-nitrophenyl ethyl phosphate | mutant enzyme Q13A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 57 | - |
p-nitrophenyl ethyl phosphate | mutant enzyme H218A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 70 | - |
p-nitrophenyl ethyl phosphate | Km above 70 mM, mutant enzyme Y105A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 73 | - |
p-nitrophenyl ethyl phosphate | mutant enzyme K337A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
Metals/Ions
| Metals/Ions | Comment | Organism | Structure |
|---|---|---|---|
| Ca2+ | contains 0.41 Ca2+ per protein molecule | Rhizobium leguminosarum | |
| Fe2+ | contains 0.17 Fe2+ per protein molecule | Rhizobium leguminosarum | |
| Mn2+ | contains 0.19 Mn2+ per protein molecule, Mn2+ is the native metal ion | Rhizobium leguminosarum | |
| Zn2+ | contains 0.22 Zn2+ per protein molecule | Rhizobium leguminosarum | |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Rhizobium leguminosarum | - |
Rhizobium leguminosarum bv. viciae strain 3841 | - |
Purification (Commentary)
| Purification (Comment) | Organism |
|---|---|
| Strep-tactin column chromatography and Superdex 200 gel filtration | Rhizobium leguminosarum |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| additional information | PMH hydrolyses phosphonate monoesters and phosphate diesters with similar efficiency | Rhizobium leguminosarum | ? | - |
? | |
| p-nitrophenyl ethyl phosphate + H2O | - |
Rhizobium leguminosarum | p-nitrophenol + ethyl hydrogen phosphate | - |
? | |
| p-nitrophenyl phenylphosphonate + H2O | - |
Rhizobium leguminosarum | p-nitrophenol + phenylphosphonic acid | - |
? | |
Subunits
| Subunits | Comment | Organism |
|---|---|---|
| tetramer | dimer of dimers in solution, X-ray crystallography | Rhizobium leguminosarum |
Synonyms
| Synonyms | Comment | Organism |
|---|---|---|
| phosphonate monoester hydrolase/phosphodiesterase | - |
Rhizobium leguminosarum |
| PMH | - |
Rhizobium leguminosarum |
Turnover Number [1/s]
| Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|
| 0.0047 | - |
p-nitrophenyl ethyl phosphate | mutant enzyme C57A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 0.0088 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme C57A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 0.17 | - |
p-nitrophenyl ethyl phosphate | kcat below 0.17 sec-1, mutant enzyme Y105A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 0.3 | - |
p-nitrophenyl ethyl phosphate | mutant enzyme H218A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 0.57 | - |
p-nitrophenyl ethyl phosphate | mutant enzyme T107A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 1 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme H218A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 1.1 | - |
p-nitrophenyl ethyl phosphate | mutant enzyme N78A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 1.5 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme N78A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 1.6 | - |
p-nitrophenyl ethyl phosphate | mutant enzyme C57S, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 2.4 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme T107A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 2.7 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme Y105A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 2.9 | - |
p-nitrophenyl ethyl phosphate | mutant enzyme K337A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 3.3 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme C57S, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 9.5 | - |
p-nitrophenyl ethyl phosphate | wild type enzyme, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 12 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme K337A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 16 | - |
p-Nitrophenyl phenylphosphonate | wild type enzyme, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 18 | - |
p-nitrophenyl ethyl phosphate | mutant enzyme Q13A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
| 26 | - |
p-Nitrophenyl phenylphosphonate | mutant enzyme Q13A, at 30°C, in 44 mM succinic acid, 33 mM imidazole, 33 mM diethanolamine, pH 7.5 | Rhizobium leguminosarum | |
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