Information on EC 1.5.1.37 - FAD reductase (NADH)

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
1.5.1.37
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RECOMMENDED NAME
GeneOntology No.
FAD reductase (NADH)
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
FADH2 + NAD+ = FAD + NADH + H+
show the reaction diagram
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-
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Riboflavin metabolism
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Metabolic pathways
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Microbial metabolism in diverse environments
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SYSTEMATIC NAME
IUBMB Comments
FADH2:NAD+ oxidoreductase
The enzyme from Burkholderia phenoliruptrix can reduce either FAD or flavin mononucleotide (FMN) but prefers FAD. Unlike EC 1.5.1.36, flavin reductase (NADH), the enzyme can not reduce riboflavin. The enzyme does not use NADPH as acceptor.
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
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
FAD + NADH + H+
FADH2 + NAD+
show the reaction diagram
FADH2 + NAD+
FAD + NADH + H+
show the reaction diagram
FMN + NADH + H+
FMNH2 + NAD+
show the reaction diagram
lumiflavin + NADH + H+
reduced lumiflavin + NAD+
show the reaction diagram
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6% of the activity with FAD
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?
riboflavin + NADH + H+
reduced riboflavin + NAD+
show the reaction diagram
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9% of the activity with FAD
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?
additional information
?
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methylene blue and ferricyanide are less than 5% as effective as FAD
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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
FAD + NADH + H+
FADH2 + NAD+
show the reaction diagram
FADH2 + NAD+
FAD + NADH + H+
show the reaction diagram
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
FMN
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12% of the activity with FAD
Lumiflavin
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6% of the activity with FAD
riboflavin
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9% of the activity with FAD
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
AMP
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AMP exhibits competitive inhibition versus NADH and noncompetitive inhibition versus FAD
NAD+
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NAD+ demonstrates competitive inhibition versus NADH and noncompetitive inhibition versus FAD
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0032 - 0.0082
FAD
0.0103 - 0.0286
FMN
0.0584
Lumiflavin
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pH 7.0, 30°C
0.0401 - 0.164
NADH
0.0383
riboflavin
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pH 7.0, 30°C
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
4.5 - 65
FAD
18.5 - 72.4
FMN
66.5
Lumiflavin
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pH 7.0, 30°C
3.1 - 62.6
NADH
68.2
riboflavin
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pH 7.0, 30°C
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
3458
FAD
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cosubstrate: NADH, pH 7.0, 24°C
1796
FMN
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cosubstrate: NADH, pH 7.0, 24°C
72.5 - 77.3
NADH
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6
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Na-phosphate buffer
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 7.6
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pH 5.0 (sodium succinate buffer): 52% of maximal activity, pH 7.6: 75% of maximal activity
6 - 8
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pH 6.0: 73% of maximal activity, pH 8.0: 81% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
22
assay at room temperature
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
23 - 45
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23°C: 85% of maximal activity, 45°C: 70% of maximal activity
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.3
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calculated from sequence
6.5
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isoelectric focusing
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
19000
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2 * 19000, calculation from sequence
19400
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deduced from cDNA, without His-tag but with FAD
22000
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x * 22000, His-tagged TftC, SDS-PAGE
39000
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gel filtration
additional information
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the apparent molecular mass of the PrnD-PrnF complex appears to be approximately 120000 Da (gel filtration), confirming a 1:1 stoichiometry for binding of the two proteins, PrnD (86000 Da) and PrnF (39000 Da)
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
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x * 22000, His-tagged TftC, SDS-PAGE
dimer
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2 * 19000, calculation from sequence
monomer
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gel filtration
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
apo-form crystals of TftC, TftC-FAD complex and TftC-FAD-NADH complex are grown at 4°C using the hanging drop vapor diffusion method. Crystal structures of dimeric TftC is determined at 2.5 A resolution
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4
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pH 7.0, 1 mM dithiothreitol, 50 mM Tris-HCl, 50% loss of activity after 12 days
16
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pH 7.0, 1 mM dithiothreitol, 50 mM Tris-HCl, 50% loss of activity after 5 days
30
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pH 7.0, 1 mM dithiothreitol, 50 mM Tris-HCl, 50% loss of activity after 1 day
42
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pH 7.0, 1 mM dithiothreitol, 50 mM Tris-HCl, 50% loss of activity after 30 min
50
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pH 7.0, 1 mM dithiothreitol, 50 mM Tris-HCl, 50% loss of activity after 1 min
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-80°C, His-tagged TftC loses about 50% activity after 2 months
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0°C, His-tagged TftC is unstable in the elution buffer containing 140 mM imidazole on ice, losing about 50% activity in 2 h
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
the reductase is purified as a stable C-terminally His-tagged yellow protein containing weakly bound FAD
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli as a His-tagged fusion protein
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expressed in Escherichia coli BL21(DE3) cells
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expression in Escherichia coli as a C-terminally His-tagged fusion protein
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overexpression in Escherichia coli
overexpression in Escherichia coli in a soluble form
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quantitative real-time PCR enzyme expression analysis
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
upregulation of the NADH:FAD oxidoreductase due to silencing of an NADH dehydrogenase
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information