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Literature summary for 4.2.1.84 extracted from
- Enhancement of thermo-stability and product tolerance of Pseudomonas putida nitrile hydratase by fusing with self-assembling peptide (2014), J. Biosci. Bioeng., 118, 249-252.
Cloned(Commentary)
| Cloned (Comment) | Organism |
|---|---|
| expression of His-tagged wild-type enzyme and peptide fused enzyme variants in Escherichia coli strain BL21(DE3) | Pseudomonas putida |
Protein Variants
| Protein Variants | Comment | Organism |
|---|---|---|
| additional information | the thermostability and product tolerance of nitrile hydratase are enhanced by fusing with two of the self-assembling amphipathic peptides, EAK16 (AEAEAKAKAEAEAKAK) at the N- and C-terminus and ELK16 (LELELKLKLELELKLK) at the N-terminus. When self-assembling amphipathic peptide ELK16 is fused to the N-terminus of the enzymes beta-subunit, the resultant enzyme (SAP-NHase-2) becomes an active inclusion body, while EAK16 does not affect the enzyme solubility when fused to the enzyme's C-terminus (SAP-NHase-10) or N-termninus (AP-NHase-1) of the beta-subunit | Pseudomonas putida |
Inhibitors
| Inhibitors | Comment | Organism | Structure |
|---|---|---|---|
| Acrylamide | NHases exhibit low product acrylamide tolerance | Pseudomonas putida |  |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Pseudomonas putida | - |
- |
- |
| Pseudomonas putida NRRL-18668 | - |
- |
- |
Purification (Commentary)
| Purification (Comment) | Organism |
|---|---|
| recombinant His-tagged wild-type enzyme and peptide fused enzyme variants from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and gel filtration | Pseudomonas putida |
Specific Activity [micromol/min/mg]
| Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
|---|---|---|---|
| 13 | - |
cell extract supernatant containing recombinant His-tagged ELK-16 N-terminal fusion enzyme variant, pH 7.5, 20°C | Pseudomonas putida |
| 109 | - |
cell extract supernatant containing recombinant His-tagged EAK-16 N-terminal fusion enzyme variant, pH 7.5, 20°C | Pseudomonas putida |
| 110 | - |
cell extract supernatant containing recombinant His-tagged wild-type enzyme, pH 7.5, 20°C | Pseudomonas putida |
| 372 | - |
purified recombinant His-tagged ELK-16 N-terminal fusion enzyme variant, pH 7.5, 20°C | Pseudomonas putida |
| 426 | - |
purified recombinant His-tagged EAK-16 N-terminal fusion enzyme variant, pH 7.5, 20°C | Pseudomonas putida |
| 429 | - |
purified recombinant His-tagged EAK-16 C-terminal fusion enzyme variant, pH 7.5, 20°C | Pseudomonas putida |
| 439 | - |
purified recombinant His-tagged wild-type enzyme, pH 7.5, 20°C | Pseudomonas putida |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| 3-cyanopyridine + H2O | - |
Pseudomonas putida | pyridine-3-carbamide | i.e. nicotinamide | ? | |
| 3-cyanopyridine + H2O | - |
Pseudomonas putida NRRL-18668 | pyridine-3-carbamide | i.e. nicotinamide | ? | |
| acrylonitrile + H2O | - |
Pseudomonas putida | acrylamide | - |
? | |
| acrylonitrile + H2O | - |
Pseudomonas putida NRRL-18668 | acrylamide | - |
? | |
Synonyms
| Synonyms | Comment | Organism |
|---|---|---|
| NHase | - |
Pseudomonas putida |
Temperature Optimum [°C]
| Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 20 | - |
assay at | Pseudomonas putida |
Temperature Stability [°C]
| Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 50 | - |
wild-type enzyme, 30 min, deactivation, enzyme fused to self-assembling amphipathic peptide ELK16 (LELELKLKLELELKLK) retains 30% activity, and SAP-NHase-1 N-terminally fused to self-assembling amphipathic peptide EAK16 (AEAEAKAKAEAEAKAK) retains 45% activity, C-terminally fused enzyme SAP-NHase-10 50% activity. Treatment with buffer containing 10% acrylamide increases the thermostability so that the wild-type retains 30% activity, and the fused enzymes SAP-NHase-1, SAP-NHase-2, and SAP-NHase-10 retain 52%, 42%, and 55% activity, respectively | Pseudomonas putida |
pH Optimum
| pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|
| 7.5 | - |
assay at | Pseudomonas putida |
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