Information on EC 1.14.13.242 - 3-hydroxy-2-methylpyridine-5-carboxylate monooxygenase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
1.14.13.242
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RECOMMENDED NAME
GeneOntology No.
3-hydroxy-2-methylpyridine-5-carboxylate monooxygenase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
3-hydroxy-2-methylpyridine-5-carboxylate + NAD(P)H + H+ + O2 = 2-(acetamidomethylene)succinate + NAD(P)+
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
oxidation
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-
-
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redox reaction
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-
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reduction
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-
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
vitamin B6 degradation
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Vitamin B6 metabolism
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Microbial metabolism in diverse environments
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SYSTEMATIC NAME
IUBMB Comments
3-hydroxy-2-methylpyridine-5-carboxylate,NAD(P)H:oxygen oxidoreductase (ring-opening)
A flavoprotein (FAD).
CAS REGISTRY NUMBER
COMMENTARY hide
37256-69-2
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GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
physiological function
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
2-methyl-3-hydroxypyridine-5-carboxylic acid + NADH + O2 + H+
2-(acetamidomethylene)succinate + NAD+
show the reaction diagram
-
-
-
?
3-hydroxy-2-methylpyridine-5-carboxylate + NAD(P)H + H+ + O2
2-(acetamidomethylene)succinate + NAD(P)+
show the reaction diagram
-
-
-
-
?
3-hydroxy-2-methylpyridine-5-carboxylate + NADH + H+ + O2
2-(acetamidomethylene)succinate + NAD+
show the reaction diagram
3-hydroxy-2-methylpyridine-5-carboxylate + NADH + H+ + O2
2-(acetamidomethylene)succinate + NAD+ + H2O
show the reaction diagram
3-hydroxy-2-methylpyridine-5-carboxylate + NADH + O2
2-(acetamidomethylene)succinate + NAD(P)+
show the reaction diagram
3-hydroxy-2-methylpyridine-5-carboxylate + NADH + O2
2-(acetamidomethylene)succinate + NAD+
show the reaction diagram
3-hydroxy-2-methylpyridine-5-carboxylate + NADH + O2 + H+
2-(acetamidomethylene)succinate + NAD+
show the reaction diagram
3-hydroxy-2-methylpyridine-5-carboxylate + NADPH + H+ + O2
2-(acetamidomethylene)succinate + NADP+
show the reaction diagram
-
-
-
-
?
5-hydroxynicotinate + NAD(P)H + H+ + O2
? + NAD(P)+
show the reaction diagram
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-
-
-
?
5-hydroxynicotinic acid + NADH + H+ + O2
alpha-(N-formylaminomethylene)succinic acid + NAD+ + H2O
show the reaction diagram
5-hydroxynicotinic acid + NADH + O2
?
show the reaction diagram
5-hydroxynicotinic acid + NADH + O2 + H+
2-(formylamidomethylene)succinate + NAD+
show the reaction diagram
5-pyridoxic acid + NADH + H+ + O2
alpha-(N-acetylaminomethylene)-beta-hydroxymethyl succinic acid + NAD+ + H2O
show the reaction diagram
5-pyridoxic acid + NADH + O2
?
show the reaction diagram
-
-
-
-
?
5-pyridoxic acid + NADH + O2 + H+
2-(acetamidomethylene)-beta-hydroxymethyl succinate + NAD+
show the reaction diagram
5% of the activity with 2-methyl-3-hydroxypyridine-5-carboxylic acid
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-
?
N-methyl-5-hydroxynicotinic acid + NADH + O2
?
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
2-methyl-3-hydroxypyridine-5-carboxylic acid + NADH + O2 + H+
2-(acetamidomethylene)succinate + NAD+
show the reaction diagram
Q988D3
-
-
-
?
3-hydroxy-2-methylpyridine-5-carboxylate + NADH + H+ + O2
2-(acetamidomethylene)succinate + NAD+
show the reaction diagram
3-hydroxy-2-methylpyridine-5-carboxylate + NADH + O2
2-(acetamidomethylene)succinate + NAD+
show the reaction diagram
3-hydroxy-2-methylpyridine-5-carboxylate + NADPH + H+ + O2
2-(acetamidomethylene)succinate + NADP+
show the reaction diagram
-
-
-
-
?
5-hydroxynicotinic acid + NADH + O2 + H+
2-(formylamidomethylene)succinate + NAD+
show the reaction diagram
Q988D3
about 100% of the activity with 2-methyl-3-hydroxypyridine-5-carboxylic acid
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-
?
5-pyridoxic acid + NADH + O2 + H+
2-(acetamidomethylene)-beta-hydroxymethyl succinate + NAD+
show the reaction diagram
Q988D3
5% of the activity with 2-methyl-3-hydroxypyridine-5-carboxylic acid
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-
?
additional information
?
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
NAD(P)H
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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no metal ion required
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1-deaza-FAD
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5-hydroxynicotinic acid
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5-pyridoxic acid
6-Methylnicotinate
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6-methylnicotinic acid
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competitive with 3-hydroxy-2-methylpyridine-5-carboxylate
NAD+
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binds competitively with O2, but not with NADH
p-chloromercuribenzoate
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; 0.05 mM, quick and complete inhibition
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0254
3-Hydroxy-2-methylpyridine-5-carboxylate
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pH 8.0, 25°C
0.045 - 0.21
5-hydroxynicotinic acid
0.00056 - 0.0011
N-methyl-5-hydroxynicotinic acid
0.0054 - 0.47
NADH
0.0059 - 0.148
O2
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
11.8
3-Hydroxy-2-methylpyridine-5-carboxylate
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pH 8.0, 25°C
0.05 - 1300
5-hydroxynicotinic acid
additional information
additional information
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kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.1 - 22000
5-hydroxynicotinic acid
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00046
1-deaza-FAD
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0.023
5-pyridoxic acid
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IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.48
5-hydroxynicotinic acid
Mesorhizobium loti;
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wild type enzyme, at pH 8.0 and 25°C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3.28
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pH 8.0, 25°C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.5 - 8
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7
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substrate 3-hydroxy-2-methylpyridine-5-carboxylate
8
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substrate 5-hydroxynicotinic acid
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20
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79% of maximum activity
40
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82% of maximum activity
PDB
SCOP
CATH
UNIPROT
ORGANISM
Mesorhizobium japonicum (strain LMG 29417 / CECT 9101 / MAFF 303099);
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
41700
4 * 41700, calculation from nucleotide sequence
43000
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4 * 43000, SDS-PAGE
160000
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equilibrium sedimentation
164000
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gel fitlration
166000
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equilibrium sedimentation
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homodimer
crystal structure, the tetramer may have been disrupted
tetramer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
density functional theory/molecular mechanics calculations show that the active-site residues Arg211 and Tyr223 have a minor effect on the reaction, while the peptide bond of Pro295-Ala296, the side chain of Tyr82 and several crystal water molecules affect the reaction energy profile considerably. The ring-opening pathway, in which an epoxy transition state is formed, is more favored than the direct C2-C3 cleavage pathway. Both the reaction barriers for the hydroxylation and the ring-opening pathways are sensitive to the quantum mechanics/molecular mechanics partitioning
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hanging drop vapor diffusion method, using 8% (w/v) PEG 8000, 0.1 M Tris-HCl, pH 8.5; structure of enzyme with and without substrates 2-methyl-3-hydroxypyridine-5-carboxylic acid, 5-hydroxynicotinic acid and 5-pyridoxic acid, and of mutant Y270F. Residue Y270 is located in the active site
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
40
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stable for 10 min
OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
the enzyme is very sensitive to oxidation, it loses activity rapidly in absence of mercaptoethanol even at 4°C, it is further stabilized in presence of high concentrations of glycerol or by serum albumin
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439056
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20°C, 50% glycerol, 0.1% 2-mercaptoethanol, 1 month
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Q-Sepharose column chromatography and G-25 Sephadex gel filtration
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
; expressed in Escherichia coli BL21(DE3) cells
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expressed in Escherichia coli BL21(DE3) cells
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expression in Escherichia coli
His-tag, expressed in Escherichia coli
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Y270A
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less than 0.1% of the pyridine ring cleavage capacity of wild-type; the mutant shows reduced ring opening activity but increased NADH oxidation activity compared to the wild type enzyme
Y270F
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about 1.5% of the pyridine ring cleavage capacity of wild-type; the mutant shows reduced ring opening activity but increased NADH oxidation activity compared to the wild type enzyme
Y223E
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the mutant shows reduced activity compared to the wild type enzyme
Y223F
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the mutant shows reduced activity compared to the wild type enzyme
Y223H
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the mutant shows reduced activity compared to the wild type enzyme
Y223T
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the mutant shows reduced activity compared to the wild type enzyme
Y82F
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the mutant shows reduced activity compared to the wild type enzyme
Y82H
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the mutant shows reduced activity compared to the wild type enzyme
Y82R
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the mutant shows reduced activity compared to the wild type enzyme