Cloned (Comment) | Organism |
---|---|
expressed in Escherichia coli JM109 cells | Pseudomonas reinekei |
Crystallization (Comment) | Organism |
---|---|
in complex with (4S)-4-methylmuconolactone and (4S)-3-methylmuconolactone, sitting drop vapor diffusion method, at 19°C, using 25% (w/v) polyethylene glycol 1500, 0.1 M MMT buffer, pH 5 (DL-malic acid, MES, and Tris base in a 1:2:2 molar ratio) | Pseudomonas reinekei |
Protein Variants | Comment | Organism |
---|---|---|
C67S | incubation of variant C67S with 4-chloromercuribenzoate does not significantly decrease its activity (above 65% of activity remaining after a 30-min incubation) | Pseudomonas reinekei |
H26A | the mutant shows a marked decrease in catalytic activity (kcat) by two orders of magnitude | Pseudomonas reinekei |
H52A | the mutation results in an increase in Km by a factor of 9, but only a small change in kcat | Pseudomonas reinekei |
Y39F | the mutant displays reduced activity compared to the wild type enzyme | Pseudomonas reinekei |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
4-chloromercuribenzoate | remaining activity of 6% after incubation with 0.02 mM 4-chloromercuribenzoate for 30 min | Pseudomonas reinekei |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.045 | - |
(4S)-4-methylmuconolactone | mutant enzyme Y39F, in phosphate buffer, 50 mM, pH 7.3 | Pseudomonas reinekei | |
0.052 | - |
(4S)-4-methylmuconolactone | wild type enzyme, in phosphate buffer, 50 mM, pH 7.3 | Pseudomonas reinekei | |
0.0972 | - |
(4S)-4-methylmuconolactone | mutant enzyme C67S, in phosphate buffer, 50 mM, pH 7.3 | Pseudomonas reinekei | |
0.4583 | - |
(4S)-4-methylmuconolactone | mutant enzyme H52A, in phosphate buffer, 50 mM, pH 7.3 | Pseudomonas reinekei |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Pseudomonas reinekei | C5MR76 | - |
- |
Purification (Comment) | Organism |
---|---|
Strep-Tactin-Sepharose column chromatography | Pseudomonas reinekei |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
28.8 | - |
toward 1-methylbis-lactone, in phosphate buffer, 50 mM, pH 7.3 | Pseudomonas reinekei |
57.3 | - |
toward (4S)-4-methylmuconolactone, in phosphate buffer, 50 mM, pH 7.3 | Pseudomonas reinekei |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
(4S)-4-methylmuconolactone | - |
Pseudomonas reinekei | (4S)-3-methylmuconolactone | - |
? | |
1-methylbis-lactone | - |
Pseudomonas reinekei | ? | - |
? | |
4-Carboxymethyl-4-methylbut-2-en-1,4-olide | - |
Pseudomonas reinekei | 4-Carboxymethyl-3-methylbut-2-en-1,4-olide | - |
? | |
additional information | activity with 3-methyl-cis,cis-muconate is below the detection limit | Pseudomonas reinekei | ? | - |
? |
Subunits | Comment | Organism |
---|---|---|
homodimer | x-ray crystallography | Pseudomonas reinekei |
Synonyms | Comment | Organism |
---|---|---|
4-Methylmuconolactone methylisomerase | - |
Pseudomonas reinekei |
MLMI | - |
Pseudomonas reinekei |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
5.8 | - |
(4S)-4-methylmuconolactone | mutant enzyme Y39F, in phosphate buffer, 50 mM, pH 7.3 | Pseudomonas reinekei | |
7.3 | - |
(4S)-4-methylmuconolactone | mutant enzyme C67S, in phosphate buffer, 50 mM, pH 7.3 | Pseudomonas reinekei | |
24.2 | - |
(4S)-4-methylmuconolactone | wild type enzyme, in phosphate buffer, 50 mM, pH 7.3 | Pseudomonas reinekei | |
47.2 | - |
(4S)-4-methylmuconolactone | mutant enzyme H52A, in phosphate buffer, 50 mM, pH 7.3 | Pseudomonas reinekei |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
6.3 | 6.5 | - |
Pseudomonas reinekei |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
100 | - |
(4S)-4-methylmuconolactone | mutant enzyme C67S, in phosphate buffer, 50 mM, pH 7.3 | Pseudomonas reinekei | |
100 | - |
(4S)-4-methylmuconolactone | mutant enzyme H52A, in phosphate buffer, 50 mM, pH 7.3 | Pseudomonas reinekei | |
100 | - |
(4S)-4-methylmuconolactone | mutant enzyme Y39F, in phosphate buffer, 50 mM, pH 7.3 | Pseudomonas reinekei | |
500 | - |
(4S)-4-methylmuconolactone | wild type enzyme, in phosphate buffer, 50 mM, pH 7.3 | Pseudomonas reinekei |