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Literature summary for 1.4.3.12 extracted from

  • Li, G.; Ren, J.; Iwaki, H.; Zhang, D.; Hasegawa, Y.; Wu, Q.; Feng, J.; Lau, P.C.; Zhu, D.
    Substrate profiling of cyclohexylamine oxidase and its mutants reveals new biocatalytic potential in deracemization of racemic amines (2014), Appl. Microbiol. Biotechnol., 98, 1681-1689.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli BL21(DE3) cells Microbacterium oxydans

Protein Variants

Protein Variants Comment Organism
L199A the mutant shows generally lower activity (decrease of 15-97%) towards most substrates compared to wild type enzyme with the exception of the larger substrates, such as cyclooctanamine and bicyclic (S)-1-aminotetraline Microbacterium oxydans
L353M the mutant shows 7-445% higher activity towards primary aliphatic amines with cycloalkane or aromatic moieties Microbacterium oxydans
M226A the mutant displays an enhanced activity (5-400%) towards most substrates Microbacterium oxydans
Y321A the mutant shows higher catalytic efficiency towards cyclooctanamine compared to the wild type enzyme Microbacterium oxydans
Y321F the mutant shows higher catalytic efficiency towards cyclooctanamine compared to the wild type enzyme Microbacterium oxydans

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.07
-
(S)-1-aminotetraline mutant enzyme Y321F, at pH 6.5 and 30°C Microbacterium oxydans
0.08
-
(S)-1-aminotetraline mutant enzyme Y321A, at pH 6.5 and 30°C Microbacterium oxydans
0.08
-
(S)-1-aminotetraline wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
0.21
-
(S)-1-aminotetraline mutant enzyme L353M, at pH 6.5 and 30°C Microbacterium oxydans
0.22
-
cyclooctanamine mutant enzyme Y321F, at pH 6.5 and 30°C Microbacterium oxydans
0.24
-
cyclooctanamine mutant enzyme Y321A, at pH 6.5 and 30°C Microbacterium oxydans
0.26
-
(S)-1-aminotetraline mutant enzyme M226A, at pH 6.5 and 30°C Microbacterium oxydans
0.32
-
(S)-1-aminotetraline mutant enzyme L199A, at pH 6.5 and 30°C Microbacterium oxydans
0.59
-
cyclooctanamine wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
1.08
-
Cyclohexylamine wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
1.89
-
cyclooctanamine mutant enzyme M226A, at pH 6.5 and 30°C Microbacterium oxydans
1.91
-
cyclooctanamine mutant enzyme L199A, at pH 6.5 and 30°C Microbacterium oxydans
2.04
-
cyclooctanamine mutant enzyme L353M, at pH 6.5 and 30°C Microbacterium oxydans
2.28
-
Cyclohexylamine mutant enzyme Y321F, at pH 6.5 and 30°C Microbacterium oxydans
3.39
-
Cyclohexylamine mutant enzyme L353M, at pH 6.5 and 30°C Microbacterium oxydans
4.76
-
Cyclohexylamine mutant enzyme M226A, at pH 6.5 and 30°C Microbacterium oxydans
5.94
-
Cyclohexylamine mutant enzyme Y321A, at pH 6.5 and 30°C Microbacterium oxydans
7.71
-
Cyclohexylamine mutant enzyme L199A, at pH 6.5 and 30°C Microbacterium oxydans

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
50000
-
x * 50000, SDS-PAGE Microbacterium oxydans

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
cyclohexylamine + O2 + H2O Microbacterium oxydans best substrate cyclohexanone + NH3 + H2O2
-
?
cyclohexylamine + O2 + H2O Microbacterium oxydans IH-35A best substrate cyclohexanone + NH3 + H2O2
-
?

Organism

Organism UniProt Comment Textmining
Microbacterium oxydans
-
-
-
Microbacterium oxydans IH-35A
-
-
-

Purification (Commentary)

Purification (Comment) Organism
DEAE column chromatography, and gel filtration Microbacterium oxydans

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
0.013
-
with 2-ethyl-1-hexylamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
0.018
-
with (R)-1-(4-chlorophenyl)ethylamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
0.08
-
with N-methylcyclohexanamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
0.084
-
with (R)-1-(4-fluorophenyl)ethanamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
0.11
-
with 4-methylbenzylamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
0.27
-
with (S)-1-p-tolylethanamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
0.33
-
with cyclopentylamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
0.34
-
with benzylamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
0.68
-
with 2-amino-4-phenylbutane as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
0.97
-
with 2-aminocyclohexanol as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
1.033
-
with cyclooctanamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
1.13
-
with 2-aminopentane as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
1.2
-
with 2-aminoheptane as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
1.49
-
with 2-aminonorbornane hydrochloride as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
1.66
-
with (S)-1-phenylpropropan-1-amine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
2.35
-
with (S)-1-aminoindan as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
2.36
-
with 1-phenylethanamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
2.62
-
with (S)-1-(4-chlorophenyl)ethylamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
2.79
-
with (S)-2-aminohexane as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
3.058
-
with 2-methylcyclohexanamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
3.37
-
with 1-(4-fluorophenyl)ethanamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
3.51
-
with (S)-(-)-1-phenylethylamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
3.6
-
with 2-amino-5-methylhexane as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
4.56
-
with (S)-1-(4-fluorophenyl)ethanamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
5.6
-
with cyclohexylamine as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
5.69
-
with (S)-1-aminotetraline as substrate, wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
(R)-1-(4-chlorophenyl)ethylamine + O2 + H2O
-
Microbacterium oxydans 1-(4-chlorophenyl)ethanone + NH3 + H2O2
-
?
(R)-1-(4-fluorophenyl)ethanamine + O2 + H2O
-
Microbacterium oxydans 1-(4-fluorophenyl)ethanone + NH3 + H2O2
-
?
(R)-1-aminotetraline + O2 + H2O
-
Microbacterium oxydans ?
-
?
(S)-1-(4-chlorophenyl)ethylamine + O2 + H2O
-
Microbacterium oxydans 1-(4-chlorophenyl)ethanone + NH3 + H2O2
-
?
(S)-1-(4-fluorophenyl)ethanamine + O2 + H2O
-
Microbacterium oxydans 1-(4-fluorophenyl)ethanone + NH3 + H2O2
-
?
(S)-1-aminoindan + O2 + H2O
-
Microbacterium oxydans ?
-
?
(S)-1-aminotetraline + O2 + H2O
-
Microbacterium oxydans ?
-
?
(S)-1-aminotetraline + O2 + H2O
-
Microbacterium oxydans tetralone + NH3 + H2O2
-
?
(S)-1-p-tolylethanamine + O2 + H2O
-
Microbacterium oxydans 1-(4-methylphenyl)ethanone + NH3 + H2O2
-
?
(S)-1-phenylethylamine + O2 + H2O
-
Microbacterium oxydans ?
-
?
(S)-1-phenylpropan-1-amine + O2 + H2O
-
Microbacterium oxydans 1-phenylpropionaldehyde + NH3 + H2O2
-
?
(S)-2-aminohexane + O2 + H2O
-
Microbacterium oxydans hexan-2-one + NH3 + H2O2
-
?
1-(4-fluorophenyl)ethanamine + O2 + H2O
-
Microbacterium oxydans 1-(4-fluorophenyl)ethanone + NH3 + H2O2
-
?
1-phenylethanamine + O2 + H2O
-
Microbacterium oxydans 1-phenylacetaldehyde + NH3 + H2O2
-
?
2-amino-4-phenylbutane + O2 + H2O
-
Microbacterium oxydans 4-phenylbutan-2-one + NH3 + H2O2
-
?
2-amino-5-methylhexane + O2 + H2O
-
Microbacterium oxydans 5-methylhexane-2-one + NH3 + H2O2
-
?
2-aminocyclohexanol + O2 + H2O
-
Microbacterium oxydans ?
-
?
2-aminoheptane + O2 + H2O
-
Microbacterium oxydans heptan-2-one + NH3 + H2O2
-
?
2-aminonorbornane + O2 + H2O
-
Microbacterium oxydans ?
-
?
2-aminopentane + O2 + H2O
-
Microbacterium oxydans pentan-2-one + NH3 + H2O2
-
?
2-ethyl-1-hexylamine + O2 + H2O
-
Microbacterium oxydans 2-ethylhexanaldehyde + NH3 + H2O2
-
?
2-methylcyclohexanamine + O2 + H2O
-
Microbacterium oxydans 2-methylcyclohexanone + NH3 + H2O2
-
?
4-methylbenzylamine + O2 + H2O
-
Microbacterium oxydans 4-methylbenzaldehyde + NH3 + H2O2
-
?
benzylamine + O2 + H2O
-
Microbacterium oxydans benzaldehyde + NH3 + H2O2
-
?
benzylamine + O2 + H2O
-
Microbacterium oxydans IH-35A benzaldehyde + NH3 + H2O2
-
?
cyclohexylamine + O2 + H2O
-
Microbacterium oxydans cyclohexanone + NH3 + H2O2
-
?
cyclohexylamine + O2 + H2O best substrate Microbacterium oxydans cyclohexanone + NH3 + H2O2
-
?
cyclohexylamine + O2 + H2O
-
Microbacterium oxydans IH-35A cyclohexanone + NH3 + H2O2
-
?
cyclohexylamine + O2 + H2O best substrate Microbacterium oxydans IH-35A cyclohexanone + NH3 + H2O2
-
?
cyclooctanamine + O2 + H2O
-
Microbacterium oxydans cyclooctanone + NH3 + H2O2
-
?
cyclopentylamine + O2 + H2O best substrate Microbacterium oxydans cyclopentanone + NH3 + H2O2
-
?
additional information deracemization of 1-aminotetraline by coupling wild type enzyme with a nonselective reducing agent NH3·BH3 is a route to norsertraline Microbacterium oxydans ?
-
?
additional information trace activity with (R)-2-aminohexane, 1-hexylamine, (R)-(-)-1-phenylethylamine, (R)-1-p-tolylethanamine, (R)-1-aminoindan, (S)-(-)-Nalpha-dimethylbenzylamine, 2-methylpiperidine, 4-methylpiperidine, 2-methyl-1,2,3,4-tetrahydroquinoline, and N,N-dimethylcyclohexylamine. No activity with 1-naphthalen-1-ylethanamine Microbacterium oxydans ?
-
?
additional information deracemization of 1-aminotetraline by coupling wild type enzyme with a nonselective reducing agent NH3·BH3 is a route to norsertraline Microbacterium oxydans IH-35A ?
-
?
additional information trace activity with (R)-2-aminohexane, 1-hexylamine, (R)-(-)-1-phenylethylamine, (R)-1-p-tolylethanamine, (R)-1-aminoindan, (S)-(-)-Nalpha-dimethylbenzylamine, 2-methylpiperidine, 4-methylpiperidine, 2-methyl-1,2,3,4-tetrahydroquinoline, and N,N-dimethylcyclohexylamine. No activity with 1-naphthalen-1-ylethanamine Microbacterium oxydans IH-35A ?
-
?
N-methylcyclohexanamine + O2 + H2O
-
Microbacterium oxydans ?
-
?

Subunits

Subunits Comment Organism
? x * 50000, SDS-PAGE Microbacterium oxydans

Synonyms

Synonyms Comment Organism
CHAO
-
Microbacterium oxydans

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
40
-
the wild type enzyme is stable at 40°C for at least 30 min, but decreases rapidly above 40°C Microbacterium oxydans

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
2.96
-
(S)-1-aminotetraline mutant enzyme Y321A, at pH 6.5 and 30°C Microbacterium oxydans
3.03
-
cyclooctanamine wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
3.32
-
(S)-1-aminotetraline mutant enzyme Y321F, at pH 6.5 and 30°C Microbacterium oxydans
3.44
-
cyclooctanamine mutant enzyme Y321A, at pH 6.5 and 30°C Microbacterium oxydans
3.79
-
cyclooctanamine mutant enzyme Y321F, at pH 6.5 and 30°C Microbacterium oxydans
4.77
-
Cyclohexylamine mutant enzyme L199A, at pH 6.5 and 30°C Microbacterium oxydans
5.72
-
(S)-1-aminotetraline wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
6.62
-
cyclooctanamine mutant enzyme L199A, at pH 6.5 and 30°C Microbacterium oxydans
7.06
-
cyclooctanamine mutant enzyme M226A, at pH 6.5 and 30°C Microbacterium oxydans
7.14
-
(S)-1-aminotetraline mutant enzyme L199A, at pH 6.5 and 30°C Microbacterium oxydans
7.68
-
cyclooctanamine mutant enzyme L353M, at pH 6.5 and 30°C Microbacterium oxydans
8.52
-
Cyclohexylamine mutant enzyme Y321A, at pH 6.5 and 30°C Microbacterium oxydans
8.54
-
(S)-1-aminotetraline mutant enzyme M226A, at pH 6.5 and 30°C Microbacterium oxydans
10.62
-
(S)-1-aminotetraline mutant enzyme L353M, at pH 6.5 and 30°C Microbacterium oxydans
10.93
-
Cyclohexylamine mutant enzyme Y321F, at pH 6.5 and 30°C Microbacterium oxydans
11.48
-
Cyclohexylamine wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
15.25
-
Cyclohexylamine mutant enzyme L353M, at pH 6.5 and 30°C Microbacterium oxydans
16.33
-
Cyclohexylamine mutant enzyme M226A, at pH 6.5 and 30°C Microbacterium oxydans

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
6.5
-
-
Microbacterium oxydans

pH Stability

pH Stability pH Stability Maximum Comment Organism
6 9 the enzyme retains more than 70% of its original activity over a broad range of pH 6.0–9.0 at 4°C for 24 h. However, stability decreases quickly in more acidic condition Microbacterium oxydans

Cofactor

Cofactor Comment Organism Structure
FAD
-
Microbacterium oxydans

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.62
-
Cyclohexylamine mutant enzyme L199A, at pH 6.5 and 30°C Microbacterium oxydans
1.43
-
Cyclohexylamine mutant enzyme Y321A, at pH 6.5 and 30°C Microbacterium oxydans
3.43
-
Cyclohexylamine mutant enzyme M226A, at pH 6.5 and 30°C Microbacterium oxydans
3.47
-
cyclooctanamine mutant enzyme L199A, at pH 6.5 and 30°C Microbacterium oxydans
3.74
-
cyclooctanamine mutant enzyme M226A, at pH 6.5 and 30°C Microbacterium oxydans
3.76
-
cyclooctanamine mutant enzyme L353M, at pH 6.5 and 30°C Microbacterium oxydans
4.49
-
Cyclohexylamine mutant enzyme L353M, at pH 6.5 and 30°C Microbacterium oxydans
4.79
-
Cyclohexylamine mutant enzyme Y321F, at pH 6.5 and 30°C Microbacterium oxydans
5.14
-
cyclooctanamine wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
10.63
-
Cyclohexylamine wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans
14.33
-
cyclooctanamine mutant enzyme Y321A, at pH 6.5 and 30°C Microbacterium oxydans
17.23
-
cyclooctanamine mutant enzyme Y321F, at pH 6.5 and 30°C Microbacterium oxydans
22.31
-
(S)-1-aminotetraline mutant enzyme L199A, at pH 6.5 and 30°C Microbacterium oxydans
32.85
-
(S)-1-aminotetraline mutant enzyme M226A, at pH 6.5 and 30°C Microbacterium oxydans
37
-
(S)-1-aminotetraline mutant enzyme Y321A, at pH 6.5 and 30°C Microbacterium oxydans
47.43
-
(S)-1-aminotetraline mutant enzyme Y321F, at pH 6.5 and 30°C Microbacterium oxydans
50.57
-
(S)-1-aminotetraline mutant enzyme L353M, at pH 6.5 and 30°C Microbacterium oxydans
71.5
-
(S)-1-aminotetraline wild type enzyme, at pH 6.5 and 30°C Microbacterium oxydans