Cloned (Comment) | Organism |
---|---|
Pseudomonas putida UW4 ACC deaminase is cloned into the pET30a (+) vector at the EcoRV/HindIII sites. All single and double mutants are constructed using a Phusion Site Directed Mutagenesis Kit. | Pseudomonas putida |
Protein Variants | Comment | Organism |
---|---|---|
E295S | by site-directed mutagenesis, the Pseudomonas putida UW4 single mutant is constructed using pET30a (+) with the full-length ACC deaminase as the template | Pseudomonas putida |
E295S/L322T | the double mutant is constructed using the E295S mutant as the template | Pseudomonas putida |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
aminooxyacetic acid | 0.005-1 mM, enzymatic activity is progressively inhibited as the aminooxyacetic acid concentration is increased | Pseudomonas putida |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.34 | - |
D-cysteine | double mutant Pseudomonas putida E295S+L322T | Pseudomonas putida |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
1-aminocyclopropane-1-carboxylate + H2O | Pseudomonas putida | activity of the mutants, immediate precursor of ethylene in plants | alpha-ketobutyrate + NH3 | - |
? | |
1-aminocyclopropane-1-carboxylate + H2O | Pseudomonas putida UW4 | activity of the mutants, immediate precursor of ethylene in plants | alpha-ketobutyrate + NH3 | - |
? | |
D-cysteine + H2O | Pseudomonas putida | - |
sulfide + NH3 + pyruvate | - |
? | |
D-cysteine + H2O | Pseudomonas putida UW4 | - |
sulfide + NH3 + pyruvate | - |
? | |
additional information | Pseudomonas putida | Site-directed mutagenesis shows that altering two amino acid residues at the same positions within the predicted active site serves to change the enzyme from D-cysteine desulfhydrase to deaminase from Pseudomonas putida UW4 the enzyme is converted into D-cysteine desulfhydrase. | ? | - |
? | |
additional information | Pseudomonas putida UW4 | Site-directed mutagenesis shows that altering two amino acid residues at the same positions within the predicted active site serves to change the enzyme from D-cysteine desulfhydrase to deaminase from Pseudomonas putida UW4 the enzyme is converted into D-cysteine desulfhydrase. | ? | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Pseudomonas putida | - |
- |
- |
Purification (Comment) | Organism |
---|---|
Recombinant protein expressed with an N-terminal 6xHis-tag is purified under native/non-denaturing conditions using Ni-NTA Superflow resin. | Pseudomonas putida |
Source Tissue | Comment | Organism | Textmining |
---|---|---|---|
cell culture | strain UW4 | Pseudomonas putida | - |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
additional information | - |
activity below detection, ACC deaminase activity in double mutant E295S/L322T | Pseudomonas putida |
0.001594 | - |
D-cysteine desulfhydrase activity in wild-type enzyme | Pseudomonas putida |
0.00819 | - |
ACC deaminase activity in mutant E295S | Pseudomonas putida |
0.0356 | - |
D-cysteine desulfhydrase activity in mutant E295S | Pseudomonas putida |
0.1475 | - |
D-cysteine desulfhydrase activity in double mutant E295S+L322T | Pseudomonas putida |
2.812 | - |
ACC deaminase activity in wild-type enzyme | Pseudomonas putida |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
1-aminocyclopropane-1-carboxylate + H2O | activity of the mutants, immediate precursor of ethylene in plants | Pseudomonas putida | alpha-ketobutyrate + NH3 | - |
? | |
1-aminocyclopropane-1-carboxylate + H2O | activity of the mutants, immediate precursor of ethylene in plants | Pseudomonas putida UW4 | alpha-ketobutyrate + NH3 | - |
? | |
D-cysteine + H2O | - |
Pseudomonas putida | sulfide + NH3 + pyruvate | - |
? | |
D-cysteine + H2O | - |
Pseudomonas putida UW4 | sulfide + NH3 + pyruvate | - |
? | |
additional information | Site-directed mutagenesis shows that altering two amino acid residues at the same positions within the predicted active site serves to change the enzyme from D-cysteine desulfhydrase to deaminase from Pseudomonas putida UW4 the enzyme is converted into D-cysteine desulfhydrase. | Pseudomonas putida | ? | - |
? | |
additional information | Site-directed mutagenesis shows that altering two amino acid residues at the same positions within the predicted active site serves to change the enzyme from D-cysteine desulfhydrase to deaminase from Pseudomonas putida UW4 the enzyme is converted into D-cysteine desulfhydrase. | Pseudomonas putida UW4 | ? | - |
? |
Synonyms | Comment | Organism |
---|---|---|
ACC deaminase | - |
Pseudomonas putida |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
37 | - |
assay at | Pseudomonas putida |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
654 | - |
D-cysteine | double mutant Pseudomonas putida E295S+L322T | Pseudomonas putida |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
8 | - |
assay at | Pseudomonas putida |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
pyridoxal 5'-phosphate | ACC deaminase binds one pyridoxal phosphate (PLP) molecule at each subunit via a conserved lysine residue. | Pseudomonas putida |