Literature summary for 1.14.13.92 extracted from

Parra, L.P.; Acevedo, J.P.; Reetz, M.T.
Directed evolution of phenylacetone monooxygenase as an active catalyst for the Baeyer-Villiger conversion of cyclohexanone to caprolactone (2015), Biotechnol. Bioeng., 112, 1354-1364 .

Search for more literature for 1.14.13.92

Application

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

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 [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/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
phenylacetone + NADPH + H+ + O2	Thermobifida fusca	-	benzyl acetate + NADP+ + H2O	-	?

Organism

Organism	UniProt	Comment	Textmining
Thermobifida fusca	Q47PU3	-	-

Substrates and Products (Substrate)

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

Synonyms	Comment	Organism
PAMO	-	Thermobifida fusca

Temperature Optimum [°C]

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 [°C]

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 [1/s]

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

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
8	-	in vitro assay at	Thermobifida fusca

Cofactor

Cofactor	Comment	Organism	Structure
NADPH	-	Thermobifida fusca

General Information

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 [mM/s]

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