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Literature summary extracted from

  • Rehdorf, J.; Zimmer, C.L.; Bornscheuer, U.T.
    Cloning, expression, characterization, and biocatalytic investigation of the 4-hydroxyacetophenone monooxygenase from Pseudomonas putida JD1 (2009), Appl. Environ. Microbiol., 75, 3106-3114.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.14.13.84 PCR-amplification, expression in Escherichia coli DH5alpha, JM109, BL21 (DE3), and Rosetta Pseudomonas putida

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.14.13.84 0.0056
-
 4-aminoacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.0381
-
 4-hydroxyacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.0885
-
 3-methoxyacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.0894
-
 propiophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.103
-
 2-acetylpyrrole 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.112
-
 trifluoroacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.121
-
 2-hydroxyacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.14
-
 4-chloroacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.199
-
 4-methoxyacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.208
-
 4-fluoroacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.241
-
 4-methylacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.244
-
 3-nitroacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.246
-
 3'-hydroxyacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.257
-
 benzaldehyde 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.287
-
 3-aminoacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.313
-
 4-Nitroacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.393
-
 acetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.953
-
 2-decanone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 4.857
-
 2,4-Pentanedione 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 18.85
-
 methyl-4-tolyl sulfide 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 21.24
-
 2-phenylpropionaldehyde 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 30.75
-
 butyrophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.14.13.84 4-hydroxyacetophenone + NADPH + H+ + O2 Pseudomonas putida and related cyclic aliphatic ketones, no activity with NADH as cofactor 4-hydroxyphenyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 4-hydroxyacetophenone + NADPH + H+ + O2 Pseudomonas putida JD1 and related cyclic aliphatic ketones, no activity with NADH as cofactor 4-hydroxyphenyl acetate + NADP+ + H2O
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
1.14.13.84 Pseudomonas putida B7T912 JD1
-
1.14.13.84 Pseudomonas putida JD1 B7T912 JD1
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.14.13.84 overexpressing Escherichia coli cells are centrifuged, washed, disrupted with a French pressure cell, centrifuged, supernatant (crude extract) purified to homogeneity in 2 chromatographic steps with Q-Sepharose and Reactive Red Pseudomonas putida

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
1.14.13.84 0.54
-
 crude cell extract, decrease of NADPH, 50 mM Tris-HCl, pH 8.0, 312.5 NADPH, air saturated, 30°C Pseudomonas putida
1.14.13.84 7.5
-
 Q-Sepharose purified, decrease of NADPH, 50 mM Tris-HCl, pH 8.0, 312.5 NADPH, air saturated, 30°C Pseudomonas putida
1.14.13.84 23.2
-
 Reactive Red purified, decrease of NADPH, 50 mM Tris-HCl, pH 8.0, 312.5 NADPH, air saturated, 30°C Pseudomonas putida

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.14.13.84 2'-hydroxyacetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 2'-hydroxyacetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida JD1 ? + NADP+ + H2O
-
 ?
1.14.13.84 2,4-pentanedione + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 2-acetylpyrrole + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 2-decanone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 2-hydroxyacetophenone + NADPH + H+ + O2
-
 Pseudomonas putida 2-hydroxyphenyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 2-hydroxyacetophenone + NADPH + H+ + O2
-
 Pseudomonas putida JD1 2-hydroxyphenyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 2-phenylpropionaldehyde + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 3'-hydroxyacetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 3-aminoacetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 3-methoxyacetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 3-nitroacetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 3-phenyl-2-butanone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 25°C Pseudomonas putida 1-phenylethyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 3-phenyl-2-pentanone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 25°C Pseudomonas putida 1-phenylpropyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 4-aminoacetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 4-chloroacetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 4-fluoroacetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 4-hydroxy-2-decanone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 25°C Pseudomonas putida 2-hydroxyoctyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 4-hydroxy-2-nonanone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 25°C Pseudomonas putida 2-hydroxyheptyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 4-hydroxy-2-octanone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 25°C Pseudomonas putida 2-hydroxyhexyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 4-hydroxy-2-undecanone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 25°C Pseudomonas putida 2-hydroxynonyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 4-hydroxy-4-phenyl-2-butanone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 25°C Pseudomonas putida 1-hydroxy-1-phenylethyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 4-hydroxyacetophenone + NADPH + H+ + O2 and related cyclic aliphatic ketones, no activity with NADH as cofactor Pseudomonas putida 4-hydroxyphenyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 4-hydroxyacetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida 4-hydroxyphenyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 4-hydroxyacetophenone + NADPH + H+ + O2 and related cyclic aliphatic ketones, no activity with NADH as cofactor Pseudomonas putida JD1 4-hydroxyphenyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 4-hydroxyacetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida JD1 4-hydroxyphenyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 4-methoxyacetophenone + NADPH + H+ + O2
-
 Pseudomonas putida 4-methoxyphenyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 4-methoxyacetophenone + NADPH + H+ + O2
-
 Pseudomonas putida JD1 4-methoxyphenyl acetate + NADP+ + H2O
-
 ?
1.14.13.84 4-methoxyacetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 4-methylacetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 4-nitroacetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 acetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 benzaldehyde + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 butyrophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 methyl-4-tolyl sulfide + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 propiophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?
1.14.13.84 trifluoroacetophenone + NADPH + H+ + O2 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida ? + NADP+ + H2O
-
 ?

Synonyms

EC Number Synonyms Comment Organism
1.14.13.84 4-hydroxyacetophenone monooxygenase
-
 Pseudomonas putida
1.14.13.84 HAPMO
-
 Pseudomonas putida

Temperature Stability [°C]

EC Number Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
1.14.13.84 25 35 highest thermal stability at 30°C, at 35°C after 30 min 47% activity is retained, at 40°C less than 1%, pH 8.0, 50 mM Tris-HCl buffer Pseudomonas putida

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.14.13.84 0.4
-
 3-methoxyacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 1
-
 3-aminoacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 1.2
-
 4-methoxyacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 1.2
-
 3-nitroacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 1.3
-
 4-fluoroacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 1.6
-
 4-chloroacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 2.2
-
 benzaldehyde 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 2.6
-
 4-Nitroacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 2.6
-
 propiophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 3.5
-
 2,4-Pentanedione 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 4.2
-
 2-decanone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 4.4
-
 4-methylacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 4.9
-
 butyrophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 8.1
-
 3'-hydroxyacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 8.2
-
 4-aminoacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 8.7
-
 2-phenylpropionaldehyde 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 9.8
-
 4-hydroxyacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 11.3
-
 2-acetylpyrrole 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 15.4
-
 trifluoroacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 16.3
-
 acetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 20
-
 methyl-4-tolyl sulfide 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 21.2
-
 2'-hydroxyacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.14.13.84 8
-
 Tris-HCl, below pH 6.0 the activity is very low Pseudomonas putida

pH Range

EC Number pH Minimum pH Maximum Comment Organism
1.14.13.84 4 9 maximum activity pH at 8.0, below pH 6.0 the activity is very low, acetate buffer (pH 4.0-5.5), phosphate buffer (pH 7.5-8.0), Tris-HCl (pH 7.5-9.0), 25°, 30 min Pseudomonas putida

Cofactor

EC Number Cofactor Comment Organism Structure
1.14.13.84 NADPH
-
 Pseudomonas putida

General Information

EC Number General Information Comment Organism
1.14.13.84 metabolism recombinant Baeyer-Villiger monooxygenases converts arylaliphatic ketones into the corresponding esters with preference for para-substitution at the aromatic ring, oxidation of aldehydes and some heteroaromatic compounds are also converted while cycloketones and open-chain ketones are not or poorly accepted as substrates, aromatic ketones such as 3-phenyl-2-butanone are oxidized with excellent enantioselectivity Pseudomonas putida

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
1.14.13.84 0.2
-
 butyrophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.4
-
 2-phenylpropionaldehyde 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 0.7
-
 2,4-Pentanedione 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 1.1
-
 methyl-4-tolyl sulfide 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 3.3
-
 3-aminoacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 4.3
-
 3-methoxyacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 4.4
-
 2-decanone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 4.9
-
 3-nitroacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 5.9
-
 4-methoxyacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 6.2
-
 4-fluoroacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 8.1
-
 benzaldehyde 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 8.4
-
 4-Nitroacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 11
-
 4-chloroacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 18.5
-
 4-methylacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 28.5
-
 propiophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 32.7
-
 3'-hydroxyacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 41.6
-
 acetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 110
-
 2-acetylpyrrole 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 138
-
 trifluoroacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 176
-
 2-hydroxyacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 257
-
 4-hydroxyacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida
1.14.13.84 1450
-
 4-aminoacetophenone 312 microM NADPH, 50 mM Tris-HCl, pH 8.0, 30°C Pseudomonas putida