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Literature summary for 1.14.13.84 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)

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

KM Value [mM]

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

Natural Substrates/ Products (Substrates)

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

Organism UniProt Comment Textmining
Pseudomonas putida B7T912 JD1
-
Pseudomonas putida JD1 B7T912 JD1
-

Purification (Commentary)

Purification (Comment) Organism
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]

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

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

Synonyms

Synonyms Comment Organism
4-hydroxyacetophenone monooxygenase
-
Pseudomonas putida
HAPMO
-
Pseudomonas putida

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
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]

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

pH Optimum

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

pH Range

pH Minimum pH Maximum Comment Organism
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

Cofactor Comment Organism Structure
NADPH
-
Pseudomonas putida

General Information

General Information Comment Organism
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]

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