Crystallization (Comment) | Organism |
---|---|
purified recombinant His-tagged wild-type enzyme and mutant K42E in complex with salicylate and pyruvate, hanging drop vapor diffusion method, mixing of 0.001 ml of 64 mg/ml wild-type protein with 0.001 ml of reservoir solution containing 0.2 M lithium sulfate, 0.1 M sodium acetate, pH 4.5, and 6% glycerol, mixing of 0.001 ml of 34 mg/ml mutant protein with 0.001 ml reservoir solution containing 0.004 M Gly-Gly, 0.100 M sodium acetate, pH 3.6, and 12% glycerol, ligands in 20fold molar excess, 25°C, 24-48 h, X-ray diffraction structure determination and analysis, modeling | Pseudomonas aeruginosa |
wild-type and mutant K42E, to 1.95 and 1.79 A resolution, respectively, in complex with salicylate and pyruvate | Pseudomonas aeruginosa |
Protein Variants | Comment | Organism |
---|---|---|
K42A | similar to wild-type, active across the entire pH-range from 4 to 9, with a constant level of activity at pH 5 and above | Pseudomonas aeruginosa |
K42A | site-directed mutagenesis of the catalytic residue | Pseudomonas aeruginosa |
K42E | no detectable activity at any pH tested | Pseudomonas aeruginosa |
K42E | site-directed mutagenesis of the catalytic residue, inactive mutant, crystal structure determination and comparison to the wild-type structure | Pseudomonas aeruginosa |
K42H | the enzyme develops a pH dependence corresponding to a loss of catalytic power upon deprotonation of the histidine. With loss of the positive charge on the K42H side chain at high pH, the enzyme retains lyase activity at about 100fold lowered catalytic efficiency but loses detectable chorismate mutase activity | Pseudomonas aeruginosa |
K42H | site-directed mutagenesis of the catalytic residue, the mutant enzyme shows 100fold lowered isochorismate lyase catalytic efficiency compared to the wild-type, but loses detectable mutase activity. It develops a pH-dependence corresponding to a loss of catalytic power upon deprotonation of the histidine. The change is not due to changes in active site architecture, but due to the difference in charge at this key site | Pseudomonas aeruginosa |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
isochorismate | Pseudomonas aeruginosa | - |
salicylate + pyruvate | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Pseudomonas aeruginosa | Q51507 | isochorismate-pyruvate lyase with additional chorismate mutase activity | - |
Pseudomonas aeruginosa | Q51507 | gene pchB | - |
Reaction | Comment | Organism | Reaction ID |
---|---|---|---|
isochorismate = salicylate + pyruvate | the enzyme achieves catalysis of both pericyclic reactions, isochorismate-pyruvate lyase and chorismate-pyruvate lyase, EC 4.1.3.40, in part by the stabilization of reactive conformations and in part by electrostatic transition-state stabilization. Both, substrate organization and electrostatic transition state stabilization, contribute to catalysis | Pseudomonas aeruginosa |
Source Tissue | Comment | Organism | Textmining |
---|
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
isochorismate | - |
Pseudomonas aeruginosa | salicylate + pyruvate | - |
? | |
isochorismate | elimination of pyruvate | Pseudomonas aeruginosa | salicylate + pyruvate | - |
? | |
additional information | PchB can also perform a nonphysiological role as a chorismate mutase albeit with considerably lower catalytic efficiency | Pseudomonas aeruginosa | ? | - |
? |
Synonyms | Comment | Organism |
---|---|---|
isochorismate-pyruvate lyase | - |
Pseudomonas aeruginosa |
PchB | - |
Pseudomonas aeruginosa |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
25 | - |
assay at | Pseudomonas aeruginosa |
pH Minimum | pH Maximum | Comment | Organism |
---|---|---|---|
4 | 9 | active across the entire pH-range from 4 to 9, with a constant level of activity at pH 5 and above | Pseudomonas aeruginosa |
4 | 9 | activity range, pH profiles of recombinant wild-type and K42 mutant enzymes, overview | Pseudomonas aeruginosa |