Application | Comment | Organism |
---|---|---|
synthesis | phenylacetone monooxygenase (PAMO) is an exceptionally robust Baeyer-Villiger monooxygenase, which makes it ideal for potential industrial applications, usage as an active catalyst for the Baeyer-Villiger conversion of cyclohexanone to caprolactone, which is important as monomer in polymer science | Thermobifida fusca |
Protein Variants | Comment | Organism |
---|---|---|
A442P | random/saturation mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 81% conversion rate | Thermobifida fusca |
A442P/ L443I/S444Q | random and site-directed mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 43% conversion rate | Thermobifida fusca |
A442P/ L443V/S444Q | random and site-directed mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 45% conversion rate | Thermobifida fusca |
A442P/L443I | random/saturation mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 45% conversion rate | Thermobifida fusca |
A442P/L443L/S444Q | random and site-directed mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 41% conversion rate | Thermobifida fusca |
A442P/L443T/S444Q | random and site-directed mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 56% conversion rate | Thermobifida fusca |
A442P/L443V | random/saturation mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 90% conversion rate | Thermobifida fusca |
A442P/L443W | random/saturation mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 74% conversion rate | Thermobifida fusca |
A442P/L443W/ S444Q | random and site-directed mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 33% conversion rate | Thermobifida fusca |
L443V | random/saturation mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 53% conversion rate | Thermobifida fusca |
L443V/S444M | random/saturation mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 53% conversion rate | Thermobifida fusca |
L443V/S444Q | random/saturation mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 40-45% conversion rate | Thermobifida fusca |
L443V/S444Q | random/saturation mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 59% conversion rate | Thermobifida fusca |
L443V/S444T | random/saturation mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 57% conversion rate | Thermobifida fusca |
additional information | directed evolution of phenylacetone monooxygenase as an active catalyst for the Baeyer-Villiger conversion of cyclohexanone to caprolactone using iterative saturation mutagenesis, mutant screening, overview. Molecular dynamics simulations and induced fit docking of wild-type and mutant enzymes with cyclohexanone. The mutants are used in the whole cell system of Escherichia coli cells | Thermobifida fusca |
Q93N/P94D/P440F | random/saturation mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at low rate | Thermobifida fusca |
S441D/A442E | random/saturation mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 73% conversion rate | Thermobifida fusca |
S441G/A442P/L443T/S444Q | site-directed mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at about 90% conversion rate | Thermobifida fusca |
S441G/A442T | random/saturation mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 48% conversion rate | Thermobifida fusca |
S441H | random/saturation mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 34% conversion rate | Thermobifida fusca |
S441H/A442P | random/saturation mutagenesis, the mutant is active with cyclohexanone, in contrast to the wild-type enzyme, and catalyzes its conversion to epsilon-caprolactone at 78% conversion rate | Thermobifida fusca |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.266 | - |
cyclohexanone | pH 8.0, 25°C, recombinant mutant S441G/A442P/L443T/S444Q | Thermobifida fusca | |
0.698 | - |
cyclohexanone | pH 8.0, 25°C, recombinant mutant A442P/L443V | Thermobifida fusca |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
phenylacetone + NADPH + H+ + O2 | Thermobifida fusca | - |
benzyl acetate + NADP+ + H2O | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Thermobifida fusca | Q47PU3 | - |
- |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
cyclohexanone + NADPH + H+ + O2 | substrate of enzyme mutants, not of wild-type, overview | Thermobifida fusca | epsilon-caprolactone + NADP+ + H2O | - |
? | |
phenylacetone + NADPH + H+ + O2 | - |
Thermobifida fusca | benzyl acetate + NADP+ + H2O | - |
? |
Synonyms | Comment | Organism |
---|---|---|
PAMO | - |
Thermobifida fusca |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
25 | - |
in vitro assay at | Thermobifida fusca |
30 | 37 | in vivo assay at | Thermobifida fusca |
Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
---|---|---|---|
57.5 | - |
Tm value for enzyme PAMO mutant A442P/L443V | Thermobifida fusca |
58.5 | - |
Tm value for enzyme PAMO mutant S441G/A442P/L443T/S444Q | Thermobifida fusca |
60.5 | - |
Tm value for wild-type enzyme PAMO | Thermobifida fusca |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.156 | - |
cyclohexanone | pH 8.0, 25°C, recombinant mutant S441G/A442P/L443T/S444Q | Thermobifida fusca | |
0.304 | - |
cyclohexanone | pH 8.0, 25°C, recombinant mutant A442P/L443V | Thermobifida fusca |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
8 | - |
in vitro assay at | Thermobifida fusca |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
NADPH | - |
Thermobifida fusca |
General Information | Comment | Organism |
---|---|---|
additional information | molecular dynamics simulations and induced fit docking of wild-type and mutant enzymes with cyclohexanone | Thermobifida fusca |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.436 | - |
cyclohexanone | pH 8.0, 25°C, recombinant mutant A442P/L443V | Thermobifida fusca | |
0.577 | - |
cyclohexanone | pH 8.0, 25°C, recombinant mutant S441G/A442P/L443T/S444Q | Thermobifida fusca |