Information on EC 1.14.14.28 - long-chain alkane monooxygenase

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The expected taxonomic range for this enzyme is: Geobacillus thermodenitrificans

EC NUMBER
COMMENTARY hide
1.14.14.28
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RECOMMENDED NAME
GeneOntology No.
long-chain alkane monooxygenase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
a long-chain alkane + FMNH2 + O2 = a long-chain primary alcohol + FMN + H2O
show the reaction diagram
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SYSTEMATIC NAME
IUBMB Comments
long-chain-alkane,FMNH2:oxygen oxidoreductase
The enzyme, characterized from the bacterium Geobacillus thermodenitrificans NG80-2, is capable of converting alkanes ranging from C15 to C36 into their corresponding primary alcohols [1,2]. The FMNH2 cofactor is provided by an FMN reductase [3].
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
metabolism
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
a long-chain alkane + FMNH2 + O2
a long-chain primary alcohol + FMN + H2O
show the reaction diagram
docosan + FMNH2 + O2
1-docosanol + FMN + H2O
show the reaction diagram
dotriacontan + FMNH2 + O2
1-dotriacontanol + FMN + H2O
show the reaction diagram
hexacosan + FMNH2 + O2
1-hexacosanol + FMN + H2O
show the reaction diagram
hexadecane + FMNH2 + O2
1-hexadecanol + FMN + H2O
show the reaction diagram
hexatriacontan + FMNH2 + O2
1-hexatriacontanol + FMN + H2O
show the reaction diagram
octacosan + FMNH2 + O2
1-octacosanol + FMN + H2O
show the reaction diagram
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?
octadecane + FMNH2 + O2
1-octadecanol + FMN + H2O
show the reaction diagram
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-
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?
pentadecane + FMNH2 + O2
1-pentadecanol + FMN + H2O
show the reaction diagram
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-
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?
tetracosan + FMNH2 + O2
1-tetracosanol + FMN + H2O
show the reaction diagram
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-
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?
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
a long-chain alkane + FMNH2 + O2
a long-chain primary alcohol + FMN + H2O
show the reaction diagram
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.4 - 11
hexadecane
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.02 - 0.073
hexadecane
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0025 - 0.0315
hexadecane
32246
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
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mutant enzyme F146R/N376I
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 8.8
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both the wild-type and the mutants are active at pH values from 6.0 to 8.8
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
60
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wild-type enzyme
65
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mutant enzyme F146R/N376I
75
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mutant enzyme F146N/N376I
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
40 - 90
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both the wild-type and the mutants are active at temperatures ranging from 40°C to 90°C
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50466
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2 * 50466, calculated from sequence
105000
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homodimer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
crystals of LadA and the LadA:FMN complex are grown using the hanging-drop vapor-diffusion method at 18°C, crystal structure of the enzyme in the apoenzyme form and its complex with FMNH2
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
60
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incubation of 4 h or less results in the loss of half of the initial activity in the wild-type and F146N/N376I mutant. Mutant enzyme F146R/N376I retains more than half of the initial activity after incubation for 12 h
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
wild-type and N-terminal His-tagged fusion proteins
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
A102D
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hydroxylation activity of purified LadA mutant on hexadecane is 2.1fold higher than that of the wild-type enzyme. Hexadecane degradation rate is 2.3fold higher than that of the wild-type enzyme. A Pseudomonas fluorescens KOB2DELTA1 strain expressing the LadA mutant grows more rapidly with hexadecane than the strain expressing wild-type LadA, confirming the enhanced activity of LadA mutant in vivo. Mutant enzyme with the same size as the wild-type LadA protein. Compared to the wild-type enzyme, the mutant enzyme utilizes a narrower spectrum of n-alkanes, including C16 to C28
A102D/F146C/L320V/N376I
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mutant enzyme completely loses the catalytic activity
A102D/F146C/N376I
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mutant enzyme completely loses the catalytic activity
A102D/L320V
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mutant enzyme completely loses the catalytic activity
A102E
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hydroxylation activity of purified LadA mutant on hexadecane is 2.2fold higher than that of the wild-type enzyme. Hexadecane degradation rate is 2.2fold higher than that of the wild-type enzyme. A Pseudomonas fluorescens KOB2DELTA1 strain expressing the LadA mutant grows more rapidly with hexadecane than the strain expressing wild-type LadA, confirming the enhanced activity of LadA mutant in vivo. Mutant enzyme with the same size as the wild-type LadA protein
C14A
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point mutation completely abolishes the catalytic activity
F146C
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mutant enzyme completely loses the catalytic activity
F146C/L320V/N376I
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mutant enzyme completely loses the catalytic activity
F146C/N376I
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hydroxylation activity of purified LadA mutant on hexadecane is 2.9fold higher than that of the wild-type enzyme. Hexadecane degradation rate is 2.9fold higher than that of the wild-type enzyme. A Pseudomonas fluorescens KOB2DELTA1 strain expressing the LadA mutant grows more rapidly with hexadecane than the strain expressing wild-type LadA, confirming the enhanced activity of LadA mutant in vivo. Mutant enzyme with the same size as the wild-type LadA protein
F146E/N376I
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hydroxylation activity of purified LadA mutant on hexadecane is 2.0fold higher than that of the wild-type enzyme. Hexadecane degradation rate is 2.7fold higher than that of the wild-type enzyme. A Pseudomonas fluorescens KOB2DELTA1 strain expressing the LadA mutant grows more rapidly with hexadecane than the strain expressing wild-type LadA, confirming the enhanced activity of LadA mutant in vivo. Mutant enzyme with the same size as the wild-type LadA protein. Compared to the wild-type enzyme, the mutant enzyme utilizes a narrower spectrum of n-alkanes, including C15 to C28
F146N/N376I
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hydroxylation activity of purified LadA mutant on hexadecane is 3.4fold higher than that of the wild-type enzyme. Hexadecane degradation rate is 3.4fold higher than that of the wild-type enzyme. A Pseudomonas fluorescens KOB2DELTA1 strain expressing the LadA mutant grows more rapidly with hexadecane than the strain expressing wild-type LadA, confirming the enhanced activity of LadA mutant in vivo. The mutant enzyme shows a shift in optimum temperature from 60°C (for the wild-type enzyme) to 75°C. Compared to the wild-type enzyme, the mutant enzyme utilizes a narrower spectrum of n-alkanes, including C15 to C28
F146Q/N376I
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hydroxylation activity of purified LadA mutant on hexadecane is 2.3fold higher than that of the wild-type enzyme. Hexadecane degradation rate is 2.3fold higher than that of the wild-type enzyme. A Pseudomonas fluorescens KOB2DELTA1 strain expressing the LadA mutant grows more rapidly with hexadecane than the strain expressing wild-type LadA, confirming the enhanced activity of LadA mutant in vivo. Mutant enzyme with the same size as the wild-type LadA protein. Compared to the wild-type enzyme, the mutant enzyme utilizes a narrower spectrum of n-alkanes, including C14 to C24
F146R/N376I
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hydroxylation activity of purified LadA mutant on hexadecane is 2.5fold higher than that of the wild-type enzyme. Hexadecane degradation rate is 2.8fold higher than that of the wild-type enzyme. A Pseudomonas fluorescens KOB2DELTA1 strain expressing the LadA mutant grows more rapidly with hexadecane than the strain expressing wild-type LadA, confirming the enhanced activity of LadA mutant in vivo. Mutant enzyme with the same size as the wild-type LadA protein. The mutant enzyme is more heat resistant than wild-type protein, with more than half of the initial activity being retained after incubation at 60°C for 12 h, compared to a 60°C incubation of 4 h or less resulting in the loss of half of the initial activity in the wild-type and F146N/N376I mutant. The mutant enzyme shows a shift in optimum temperature from 60°C (for the wild-type enzyme) to 65°C. Compared to the wild-type enzyme, the mutant enzyme utilizes a narrower spectrum of n-alkanes, including C15 to C24
H17F
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the mutant maintains the same dimeric form as the wild type, point mutation completely abolishes the catalytic activity
H311F
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the mutant maintains the same dimeric form as the wild type, point mutation completely abolishes the catalytic activity
L320A
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hydroxylation activity of purified LadA mutant on hexadecane is 2.2fold higher than that of the wild-type enzyme. Hexadecane degradation rate is 2.5fold higher than that of the wild-type enzyme. A Pseudomonas fluorescens KOB2DELTA1 strain expressing the LadA mutant grows more rapidly with hexadecane than the strain expressing wild-type LadA, confirming the enhanced activity of LadA mutant in vivo. Mutant enzyme with the same size as the wild-type LadA protein. Compared to the wild-type enzyme, the mutant enzyme utilizes a narrower spectrum of n-alkanes, including C15 to C22
L320V
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hydroxylation activity of purified LadA mutant on hexadecane is 2.5fold higher than that of the wild-type enzyme. Hexadecane degradation rate is 2.4fold higher than that of the wild-type enzyme. A Pseudomonas fluorescens KOB2DELTA1 strain expressing the LadA mutant grows more rapidly with hexadecane than the strain expressing wild-type LadA, confirming the enhanced activity of LadA mutant in vivo. Mutant enzyme with the same size as the wild-type LadA protein
N376I
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mutant enzyme completely loses the catalytic activity
Q79L
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the mutant maintains the same dimeric form as the wild type, point mutation completely abolishes the catalytic activity
T63F
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the mutant maintains the same dimeric form as the wild type, point mutation completely abolishes the catalytic activity
A102D
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hydroxylation activity of purified LadA mutant on hexadecane is 2.1fold higher than that of the wild-type enzyme. Hexadecane degradation rate is 2.3fold higher than that of the wild-type enzyme. A Pseudomonas fluorescens KOB2DELTA1 strain expressing the LadA mutant grows more rapidly with hexadecane than the strain expressing wild-type LadA, confirming the enhanced activity of LadA mutant in vivo. Mutant enzyme with the same size as the wild-type LadA protein. Compared to the wild-type enzyme, the mutant enzyme utilizes a narrower spectrum of n-alkanes, including C16 to C28
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A102E
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hydroxylation activity of purified LadA mutant on hexadecane is 2.2fold higher than that of the wild-type enzyme. Hexadecane degradation rate is 2.2fold higher than that of the wild-type enzyme. A Pseudomonas fluorescens KOB2DELTA1 strain expressing the LadA mutant grows more rapidly with hexadecane than the strain expressing wild-type LadA, confirming the enhanced activity of LadA mutant in vivo. Mutant enzyme with the same size as the wild-type LadA protein
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C14A
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point mutation completely abolishes the catalytic activity
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F146C
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mutant enzyme completely loses the catalytic activity
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H17F
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the mutant maintains the same dimeric form as the wild type, point mutation completely abolishes the catalytic activity
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H311F
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the mutant maintains the same dimeric form as the wild type, point mutation completely abolishes the catalytic activity
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L320A
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hydroxylation activity of purified LadA mutant on hexadecane is 2.2fold higher than that of the wild-type enzyme. Hexadecane degradation rate is 2.5fold higher than that of the wild-type enzyme. A Pseudomonas fluorescens KOB2DELTA1 strain expressing the LadA mutant grows more rapidly with hexadecane than the strain expressing wild-type LadA, confirming the enhanced activity of LadA mutant in vivo. Mutant enzyme with the same size as the wild-type LadA protein. Compared to the wild-type enzyme, the mutant enzyme utilizes a narrower spectrum of n-alkanes, including C15 to C22
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L320V
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hydroxylation activity of purified LadA mutant on hexadecane is 2.5fold higher than that of the wild-type enzyme. Hexadecane degradation rate is 2.4fold higher than that of the wild-type enzyme. A Pseudomonas fluorescens KOB2DELTA1 strain expressing the LadA mutant grows more rapidly with hexadecane than the strain expressing wild-type LadA, confirming the enhanced activity of LadA mutant in vivo. Mutant enzyme with the same size as the wild-type LadA protein
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Q79L
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the mutant maintains the same dimeric form as the wild type, point mutation completely abolishes the catalytic activity
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T63F
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the mutant maintains the same dimeric form as the wild type, point mutation completely abolishes the catalytic activity
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
environmental protection
synthesis