Information on EC 1.5.99.4 - nicotine dehydrogenase

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The expected taxonomic range for this enzyme is: Bacteria, Eukaryota

EC NUMBER
COMMENTARY hide
1.5.99.4
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RECOMMENDED NAME
GeneOntology No.
nicotine dehydrogenase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
(S)-nicotine + acceptor + H2O = (S)-6-hydroxynicotine + reduced acceptor
show the reaction diagram
; A metalloprotein (FMN). Acts on both (R)- and (S)-isomers
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
oxidation
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redox reaction
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reduction
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Microbial metabolism in diverse environments
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Nicotinate and nicotinamide metabolism
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nicotine degradation I (pyridine pathway)
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nicotine degradation III (VPP pathway)
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SYSTEMATIC NAME
IUBMB Comments
nicotine:acceptor 6-oxidoreductase (hydroxylating)
A metalloprotein (FMN). The enzyme can act on both the naturally found (S)-enantiomer and the synthetic (R)-enantiomer of nicotine, with retention of configuration in both cases [4].
CAS REGISTRY NUMBER
COMMENTARY hide
37256-31-8
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
no activity in Escherichia coli strain DH5alpha
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Manually annotated by BRENDA team
no activity in Pseudomonas putida strain KT2440
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Manually annotated by BRENDA team
strain strain JS614
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Manually annotated by BRENDA team
male, adult, New Zealand White
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-
Manually annotated by BRENDA team
Chester
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Manually annotated by BRENDA team
Chester
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-
Manually annotated by BRENDA team
gene nox
UniProt
Manually annotated by BRENDA team
gene nox
UniProt
Manually annotated by BRENDA team
seven-week-old, male, Wistar
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-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
metabolism
physiological function
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
(R)-nicotine + acceptor + H2O
(R)-6-hydroxynicotine + reduced acceptor
show the reaction diagram
-
-
-
?
(RS)-nicotine + acceptor + H2O
pseudooxynicotine + reduced acceptor
show the reaction diagram
(S)-nicotine + acceptor + H2O
(S)-nicotine delta1',5'-iminium ion + reduced acceptor
show the reaction diagram
-
-
-
?
(S)-nicotine + acceptor + H2O
nicotine-N-oxide + reduced acceptor
show the reaction diagram
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-
-
?
(S)-nicotine + acceptor + H2O
nornicotine + reduced acceptor
show the reaction diagram
-
-
-
?
(S)-nicotine + acceptor + H2O
pseudooxynicotine + reduced acceptor
show the reaction diagram
anabasine + acceptor + H2O
6-hydroxyanabasine + reduced acceptor
show the reaction diagram
-
-
-
?
D-nicotine + acceptor + H2O
(R)-6-hydroxynicotine + reduced acceptor
show the reaction diagram
isozyme specific for the D-isomer
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-
?
DL-nicotine + ?
6-hydroxynicotine + ?
show the reaction diagram
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first enzyme in nicotine pathway
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?
L-nicotine + acceptor + H2O
(S)-6-hydroxynicotine + reduced acceptor
show the reaction diagram
myosmine + acceptor + H2O
? + reduced acceptor
show the reaction diagram
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-
-
?
nicotine + acceptor + H2O
6-hydroxynicotine + reduced acceptor
show the reaction diagram
nicotine-N-oxide + acceptor + H2O
6-hydroxynicotine-N-oxide + reduced acceptor
show the reaction diagram
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-
-
?
nicotinic acid + acceptor + H2O
6-hydroxynicotinic acid + reduced acceptor
show the reaction diagram
nornicotine + acceptor + H2O
6-hydroxynornicotine + reduced acceptor
show the reaction diagram
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-
-
?
additional information
?
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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
(R)-nicotine + acceptor + H2O
(R)-6-hydroxynicotine + reduced acceptor
show the reaction diagram
Q59127, Q59128, Q59129
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-
-
?
(S)-nicotine + acceptor + H2O
pseudooxynicotine + reduced acceptor
show the reaction diagram
D-nicotine + acceptor + H2O
(R)-6-hydroxynicotine + reduced acceptor
show the reaction diagram
Q59127, Q59128, Q59129
isozyme specific for the D-isomer
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-
?
DL-nicotine + ?
6-hydroxynicotine + ?
show the reaction diagram
-
first enzyme in nicotine pathway
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?
L-nicotine + acceptor + H2O
(S)-6-hydroxynicotine + reduced acceptor
show the reaction diagram
Q59127, Q59128, Q59129
isozyme strictly specific for the L-isomer
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-
?
nicotine + acceptor + H2O
6-hydroxynicotine + reduced acceptor
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
cytochrome c
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cytochrome P450
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FMN
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metalloflavoprotein, protects enzyme against inhibition by acriflavine
iron-sulfur centre
subunit B carries an iron-sulfur cluster
molybdenum dinucleotide cofactor
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molybdopterin cytosine dinucleotide
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pterin
additional information
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Cu2+
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inhibitory
Iron
subunit B carries an iron-sulfur cluster
Molybdenum
Tungsten
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enzyme synthesis responds to the presence of this metal ion
additional information
-
two labile sulfur groups per mol
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1,10-phenanthroline
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47% inhibition at 2 mM
2,2'-dipyridyl
8-hydroxyquinoline
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77% inhibition at 5 mM
Acriflavine
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64% inhibition at 1 mM
brilliant cresyl blue
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concentration over 0.2 mM
cyanide
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95% inhibition at 1 mM potassium cyanide
Hg2+
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96% inhibition at 0.1 mM
hydroxylamine
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22% inhibition at 5 mM
iodoacetamide
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23% inhibition at 10 mM
p-Chloromercuriphenylsulfonic acid
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62% inhibition at 1 mM
p-hydroxymercuribenzoate
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92% inhibition at 1 mM
Quinacrine
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49% inhibition at 0.1 mM
Tungsten
inhibitory to enzyme activity, does not inhibit enzyme synthesis
additional information
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
EDTA
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stimulates slightly
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.035
(S)-nicotine
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1.24
2,6-dichlorophenolindophenol
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0.008
brilliant cresyl blue
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0.49
menadione
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0.027
methylene blue
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0.037
Nicotine
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additional information
additional information
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.468 - 0.597
(S)-nicotine
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.000514 - 0.000888
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variation caused by pre-treatment with 3-methylcholanthrene, beta-naphthoflavone or phenobarbital
0.0065
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respiratory microsomes
0.0222
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olfactory microsomes
1096
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reduction of 2,6-dichlorophenolindophenol
additional information
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0.119 units per mg protein, unit is defined as the amount causing a decrease in absorbancy at 578 nm of 0.001 per min
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.4
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assay at
7.6 - 8.2
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sodium diphosphate buffer
8.6 - 9
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Tris chloride or Tris phosphate buffer
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
present both as soluble and membrane-associated form
Manually annotated by BRENDA team
present both as soluble and membrane-associated form
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Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
14924
x * 30011, subunit A, x * 14924, subunit B, x * 87677, subunit C, calculated
15100
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1 * 82000 + 1 * 30000 + 1 * 15100, SDS-PAGE
30000
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1 * 82000 + 1 * 30000 + 1 * 15100, SDS-PAGE
30011
x * 30011, subunit A, x * 14924, subunit B, x * 87677, subunit C, calculated
82000
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1 * 82000 + 1 * 30000 + 1 * 15100, SDS-PAGE
87677
x * 30011, subunit A, x * 14924, subunit B, x * 87677, subunit C, calculated
120000
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gel filtration
290000
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native gel electrophoresis
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
x * 30011, subunit A, x * 14924, subunit B, x * 87677, subunit C, calculated
trimer
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1 * 82000 + 1 * 30000 + 1 * 15100, SDS-PAGE
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-19°C, 1 month, without thawing and refreezing, without loss of activity
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-19°C, cells, 4 months, without loss of activity
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4°C, 2-3 days, no significant loss of activity
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room temperature, EDTA prevents gradual loss of activity
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate precipitation, heat denaturation, TEAE-cellulose chromatography
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ammonium sulfate precipitation, ion-exchange, gel filtration
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ammonium sulfate precipitation, ion-exchange, gel filtration; hydroxylapatite column
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protamine sulfate and ammonium sulfate precipitation, gel filtration, ion-exchange
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
gene 6hdno of pAO1, DNA and amino acid sequence determination and analysis, sequence comparisons, genetic organization
gene 6hdno of pAO1, DNA sequence analysis, genetic organization, primer extension analysis, transcriptional analysis, binding and regulatory role of repressor HdnoR, effector protein IR1 is involved, overview
gene nox, DNA and amino acid sequence deteremination and analysis, recombinant expression from a broad-host-range cloning vector in Escherichia coli strain DH5alpha and Pseudomonas putida strain KT2440, the enzyme is exxpressed in both hosts but only active in Pseudomonas putida
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
expression of all subunits requires the presence of nicotine and molybdenum in the culture medium
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information