Phenylacetone monooxygenase from Thermobifida fusca
Enzyme Description
Species
Extremophile
Yes
thermophilic organism
EC Number
1.14.13.92 β,
1.14.13.92 β (sulfoxidation)
Sequence
MAGQTTVDSRRQPPEEVDVLVVGAGFSGLYALYRLRELGRSVHVIETAGDVGGVWYWNRYPGARCDIESIEYCYSFSEEVLQEWNWTERYASQPEILRYINFVADKFDLRSGITFHTTVTAAAFDEATNTWTVDTNHGDRIRARYLIMASGQLSVPQLPNFPGLKDFAGNLYHTGNWPHEPVDFSGQRVGVIGTGSSGIQVSPQIAKQAAELFVFQRTPHFAVPARNAPLDPEFLADLKKRYAEFREESRNTPGGTHRYQGPKSALEVSDEELVETLERYWQEGGPDILAAYRDILRDRDANERVAEFIRNKIRNTVRDPEVAERLVPKGYPFGTKRLILEIDYYEMFNRDNVHLVDTLSAPIETITPRGVRTSEREYELDSLVLATGFDALTGALFKIDIRGVGNVALKEKWAAGPRTYLGLSTAGFPNLFFIAGPGSPSALSNMLVSIEQHVEWVTDHIAYMFKNGLTRSEAVLEKEDEWVEHVNEIADETLYPMTASWYTGANVPGKPRVFMLYVGGFHRYRQICDEVAAKGYEGFVLT
Francesco Secundo et al. (2010)
β Effects of water miscible organic solvents on the activity and conformation of the baeyerβvilliger monooxygenases from Thermobifida fusca and Acinetobacter calcoaceticus : A comparative study
Biotechnology and Bioengineering
Β· doi:10.1002/bit.22963 β
Β· Activity - Classical Stability - C50 Activity + Stability - Incubation
▼
Database ID
UniProt: Q47PU3 β
Sequence Annotation
Inferred - from protein name
Protein Source
Recombinant, host bacterium Escherichia coli BL21
Experimental Data (39 measurements)
39 measurements β page 2 of 2
| Property | Assay | Solvent | Solvent Volume | Aqueous Reference | Measured Value | Units | Solution | pH | Temperature | Substrate(s) | Product(s) | Cofactor(s) | Shaking | Comments |
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Activity - Classical | Activity (in U/mg) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Acetonitrile | 50% | 0.1 | 0.005 | U/mg | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Classical aqueous control (in U/mg) |
| Activity - Classical | Activity (in U/mg) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Acetonitrile | 60% | 0.1 | 0.004 | U/mg | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Classical aqueous control (in U/mg) |
| Activity - Classical | Activity (in U/mg) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Methanol | 10% | 0.1 | 0.28 | U/mg | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Classical aqueous control (in U/mg) |
| Activity - Classical | Activity (in U/mg) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Methanol | 20% | 0.1 | 0.46 | U/mg | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Classical aqueous control (in U/mg) |
| Activity - Classical | Activity (in U/mg) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Methanol | 30% | 0.1 | 0.25 | U/mg | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Classical aqueous control (in U/mg) |
| Activity - Classical | Activity (in U/mg) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Methanol | 40% | 0.1 | 0.15 | U/mg | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Classical aqueous control (in U/mg) |
| Activity - Classical | Activity (in U/mg) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Methanol | 50% | 0.1 | 0.06 | U/mg | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Classical aqueous control (in U/mg) |
| Activity - Classical | Activity (in U/mg) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Methanol | 60% | 0.1 | 0.02 | U/mg | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Classical aqueous control (in U/mg) |
| Activity - Classical | Activity (in U/mg) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Ethanol | 10% | 0.1 | 0.011 | U/mg | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Classical aqueous control (in U/mg) |
| Activity - Classical | Activity (in U/mg) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Ethanol | 20% | 0.1 | 0.01 | U/mg | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Classical aqueous control (in U/mg) |
| Activity - Classical | Activity (in U/mg) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Ethanol | 30% | 0.1 | 0.09 | U/mg | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Classical aqueous control (in U/mg) |
| Activity - Classical | Activity (in U/mg) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Ethanol | 50% | 0.1 | 0.02 | U/mg | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Classical aqueous control (in U/mg) |
| Activity + Stability - Incubation | Activity after incubation (24 hours at 25Β°C) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Methanol | 40% | 0.088 | 0.004 | U/mg | Incubation: 50 mM Tris-HCl, Assay: 50 mM Tris-HCl | Incubation: 8.6, Assay: 8.6 | Incubation: 25Β°C, Assay: 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Water-incubated control in water (in U/mg), assay in organic solvent |
| Activity + Stability - Incubation | Activity after incubation (24 hours at 25Β°C) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Methanol | 20% | 0.088 | 0.43 | U/mg | Incubation: 50 mM Tris-HCl, Assay: 50 mM Tris-HCl | Incubation: 8.6, Assay: 8.6 | Incubation: 25Β°C, Assay: 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Water-incubated control in water (in U/mg), assay in organic solvent |
| Stability - C50 | Concentration (in % v/v) for which 50% of enzyme activity is lost, as measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) | Methanol | β | β | 55.0 | % (v/v) | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Not applicable control (in % (v/v)) |
| Stability - C50 | Concentration (in % v/v) for which 50% of enzyme activity is lost, as measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) | Ethanol | β | β | 43.0 | % (v/v) | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Not applicable control (in % (v/v)) |
| Stability - C50 | Concentration (in % v/v) for which 50% of enzyme activity is lost, as measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) | 1,4-Dioxane | β | β | 26.0 | % (v/v) | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Not applicable control (in % (v/v)) |
| Stability - C50 | Concentration (in % v/v) for which 50% of enzyme activity is lost, as measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) | Acetonitrile | β | β | 22.0 | % (v/v) | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Not applicable control (in % (v/v)) |
| Stability - C50 | Concentration (in % v/v) for which 50% of enzyme activity is lost, as measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) | 2,2,2-Trifluoroethanol (TFE) | β | β | 16.0 | % (v/v) | 50 mM Tris-HCl | 8.6 | 25Β°C | Molecular oxygen (O2), Thioanisole | H2O , methyl phenyl sulfoxide | 0.1 mM NADPH | β | Not applicable control (in % (v/v)) |
Visualization : Activity β Classical
Francesco Secundo et al. (2010)
One bar per measurement. Colour = solvent, shade = solvent volume.
Visualization : Activity + Stability β Incubation
Francesco Secundo et al. (2010)
One bar per measurement. Colour = solvent, shade = solvent volume.
Visualization : Stability β C50
Francesco Secundo et al. (2010)
One bar per measurement. Colour = solvent, shade = temperature. Hover for details.
Gonzalo de Gonzalo et al. (2012)
β Improvement of the biocatalytic properties of one phenylacetone monooxygenase mutant in hydrophilic organic solvents
Enzyme and Microbial Technology
Β· doi:10.1016/j.enzmictec.2011.09.006 β
Β· Activity - Classical Stability - Incubation Activity + Stability - Conversion Specificity - Enantioselectivity
▼
Database ID
UniProt: Q47PU3 β
Sequence Annotation
Inferred - from protein name (1 mutation)
Protein Source
Recombinant, host bacterium Escherichia coli BL21
Experimental Data (67 measurements)
67 measurements β page 1 of 4
| Mutation | Property | Assay | Solvent | Solvent Volume | Aqueous Reference | WT Reference | Measured Value | Units | Solution | pH | Temperature | Substrate(s) | Product(s) | Cofactor(s) | Shaking | Comments |
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| M446G | Activity - Classical | Activity (k_obs in s^-1) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Methanol | 30% | 2.9 | β | 1.5 | 1/s | 50 mM Tris-HCl | 9 | 30Β°C | Benzyl methyl sulfide, Oβ | (S)-Benzyl dimethyl sulfoxonium | 0.2 mM NADPH | β | Classical aqueous control (in 1/s) |
| M446G | Activity - Classical | Activity (k_obs in s^-1) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Isopropanol | 10% | 2.9 | β | 2.2 | 1/s | 50 mM Tris-HCl | 9 | 30Β°C | Benzyl methyl sulfide, Oβ | (S)-Benzyl dimethyl sulfoxonium | 0.2 mM NADPH | β | Classical aqueous control (in 1/s) |
| M446G | Activity - Classical | Activity (k_obs in s^-1) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Isopropanol | 30% | 2.9 | β | 0.24 | 1/s | 50 mM Tris-HCl | 9 | 30Β°C | Benzyl methyl sulfide, Oβ | (S)-Benzyl dimethyl sulfoxonium | 0.2 mM NADPH | β | Classical aqueous control (in 1/s) |
| M446G | Activity - Classical | Activity (k_obs in s^-1) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Acetonitrile | 5% | 2.9 | β | 2.7 | 1/s | 50 mM Tris-HCl | 9 | 30Β°C | Benzyl methyl sulfide, Oβ | (S)-Benzyl dimethyl sulfoxonium | 0.2 mM NADPH | β | Classical aqueous control (in 1/s) |
| M446G | Activity - Classical | Activity (k_obs in s^-1) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Acetonitrile | 10% | 2.9 | β | 0.66 | 1/s | 50 mM Tris-HCl | 9 | 30Β°C | Benzyl methyl sulfide, Oβ | (S)-Benzyl dimethyl sulfoxonium | 0.2 mM NADPH | β | Classical aqueous control (in 1/s) |
| M446G | Activity - Classical | Activity (k_obs in s^-1) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Acetonitrile | 30% | 2.9 | β | 0.17 | 1/s | 50 mM Tris-HCl | 9 | 30Β°C | Benzyl methyl sulfide, Oβ | (S)-Benzyl dimethyl sulfoxonium | 0.2 mM NADPH | β | Classical aqueous control (in 1/s) |
| M446G | Activity - Classical | Activity (k_obs in s^-1) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Isopropanol | 5% | 2.9 | β | 2.5 | 1/s | 50 mM Tris-HCl | 9 | 30Β°C | Benzyl methyl sulfide, Oβ | (S)-Benzyl dimethyl sulfoxonium | 0.2 mM NADPH | β | Classical aqueous control (in 1/s) |
| M446G | Activity - Classical | Activity (k_obs in s^-1) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | PEG | 50% | 2.9 | β | 0.21 | 1/s | 50 mM Tris-HCl | 9 | 30Β°C | Benzyl methyl sulfide, Oβ | (S)-Benzyl dimethyl sulfoxonium | 0.2 mM NADPH | β | Classical aqueous control (in 1/s) |
| M446G | Activity - Classical | Activity (k_obs in s^-1) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | PEG | 30% | 2.9 | β | 2.1 | 1/s | 50 mM Tris-HCl | 9 | 30Β°C | Benzyl methyl sulfide, Oβ | (S)-Benzyl dimethyl sulfoxonium | 0.2 mM NADPH | β | Classical aqueous control (in 1/s) |
| M446G | Activity - Classical | Activity (k_obs in s^-1) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | PEG | 10% | 2.9 | β | 3.0 | 1/s | 50 mM Tris-HCl | 9 | 30Β°C | Benzyl methyl sulfide, Oβ | (S)-Benzyl dimethyl sulfoxonium | 0.2 mM NADPH | β | Classical aqueous control (in 1/s) |
| M446G | Activity - Classical | Activity (k_obs in s^-1) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | PEG | 5% | 2.9 | β | 3.2 | 1/s | 50 mM Tris-HCl | 9 | 30Β°C | Benzyl methyl sulfide, Oβ | (S)-Benzyl dimethyl sulfoxonium | 0.2 mM NADPH | β | Classical aqueous control (in 1/s) |
| M446G | Activity - Classical | Activity (k_obs in s^-1) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Methanol | 10% | 2.9 | β | 2.9 | 1/s | 50 mM Tris-HCl | 9 | 30Β°C | Benzyl methyl sulfide, Oβ | (S)-Benzyl dimethyl sulfoxonium | 0.2 mM NADPH | β | Classical aqueous control (in 1/s) |
| M446G | Activity - Classical | Activity (k_obs in s^-1) measured by absorbance spectrophotometry (NADPH absorbance measurement, 340 nm) in the presence of organic solvent | Methanol | 5% | 2.9 | β | 3.25 | 1/s | 50 mM Tris-HCl | 9 | 30Β°C | Benzyl methyl sulfide, Oβ | (S)-Benzyl dimethyl sulfoxonium | 0.2 mM NADPH | β | Classical aqueous control (in 1/s) |
| M446G | Activity + Stability - Conversion | Conversion after incubation (24 hours at 30Β°C) in the presence of organic solvent, measured by gas chromatography | Methanol | 10% | 81.1 | β | 97.9 | % | 50 mM Tris-HCl, 20 mM G6P, 5U G6PDH | 9 | 30Β°C | Benzyl methyl sulfide, Oβ | (S)-Benzyl dimethyl sulfoxonium | 0.2 mM NADPH | 250 rpm | Classical aqueous control (in %) |
| M446G | Activity + Stability - Conversion | Conversion after incubation (24 hours at 30Β°C) in the presence of organic solvent, measured by gas chromatography | PEG | 10% | 81.1 | β | 94.7 | % | 50 mM Tris-HCl, 20 mM G6P, 5U G6PDH | 9 | 30Β°C | Benzyl methyl sulfide, Oβ | (S)-Benzyl dimethyl sulfoxonium | 0.2 mM NADPH | 250 rpm | Classical aqueous control (in %) |
| M446G | Activity + Stability - Conversion | Conversion after incubation (24 hours at 30Β°C) in the presence of organic solvent, measured by gas chromatography | Methanol | 5% | 78.1 | β | 96.2 | % | 50 mM Tris-HCl, 20 mM G6P, 5U G6PDH | 9 | 30Β°C | Oβ, 10 mM Thioanisole | (S)-methyl phenyl sulfoxide | 0.2 mM NADPH | 250 rpm | Classical aqueous control (in %) |
| M446G | Activity + Stability - Conversion | Conversion after incubation (24 hours at 30Β°C) in the presence of organic solvent, measured by gas chromatography | Methanol | 5% | 34.9 | β | 70.2 | % | 50 mM Tris-HCl, 20 mM G6P, 5U G6PDH | 9 | 30Β°C | Oβ, 10 mM para-nitrophenyl methyl sulfide | (S)-para-nitrophenyl methyl sulfoxide | 0.2 mM NADPH | 250 rpm | Classical aqueous control (in %) |
| M446G | Activity + Stability - Conversion | Conversion after incubation (24 hours at 30Β°C) in the presence of organic solvent, measured by gas chromatography | Methanol | 5% | 90.9 | β | 98.4 | % | 50 mM Tris-HCl, 20 mM G6P, 5U G6PDH | 9 | 30Β°C | Oβ, 10 mM Phenyl methyl sulfide | (S)-Phenyl methyl sulfoxide | 0.2 mM NADPH | 250 rpm | Classical aqueous control (in %) |
| M446G | Activity + Stability - Conversion | Conversion after incubation (24 hours at 30Β°C) in the presence of organic solvent, measured by gas chromatography | Methanol | 5% | 62.4 | β | 96.4 | % | 50 mM Tris-HCl, 20 mM G6P, 5U G6PDH | 9 | 30Β°C | 10 mM Furfuryl methyl sulfide, Oβ | (S)-Furfuryl methyl sulfoxide | 0.2 mM NADPH | 250 rpm | Classical aqueous control (in %) |
| M446G | Activity + Stability - Conversion | Conversion after incubation (24 hours at 30Β°C) in the presence of organic solvent, measured by gas chromatography | Methanol | 5% | 68.9 | β | 71.2 | % | 50 mM Tris-HCl, 20 mM G6P, 5U G6PDH | 9 | 30Β°C | 10 mM Cyclohexyl methyl sulfide, Oβ | (S)-Cyclohexyl methyl sulfoxide | 0.2 mM NADPH | 250 rpm | Classical aqueous control (in %) |
Mutations in this dataset (1) β Gonzalo de Gonzalo et al. (2012)
M446G
Visualization : Activity β Classical
Gonzalo de Gonzalo et al. (2012)
One bar per measurement. Colour = solvent, shade = solvent volume.
Visualization : Stability β Incubation
Gonzalo de Gonzalo et al. (2012)
One bar per measurement. Colour = solvent, shade = solvent volume.
Visualization : Activity + Stability β Conversion
Gonzalo de Gonzalo et al. (2012)
One bar per measurement. Colour = solvent, shade = solvent volume.
Visualization : Specificity β Enantioselectivity
Gonzalo de Gonzalo et al. (2012)
One bar per measurement. Colour = solvent, shade = solvent volume.