Information on EC 1.5.1.B1 - flavin:NADH reductase

Word Map on EC 1.5.1.B1
Please wait a moment until all data is loaded. This message will disappear when all data is loaded.
Specify your search results
Select one or more organisms in this record:
Show additional data
Do not include text mining results
Include (text mining) results (more...)
Include results (AMENDA + additional results, but less precise; more...)


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

EC NUMBER
COMMENTARY hide
1.5.1.B1
preliminary BRENDA-supplied EC number
RECOMMENDED NAME
GeneOntology No.
flavin:NADH reductase
-
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
FAD + NADH + H+ = FADH2 + NAD+
show the reaction diagram
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
FAD:NADH oxidoreductase
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain W, ATCC 11105, genes hpaC and fre encoding the NAD(P)H-flavin oxidoreductase HpaC and the FAD reductase Fre
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
FAD + NAD(P)H
FADH2 + NAD(P)+
show the reaction diagram
-
-
-
-
?
FAD + NADH + H+
FADH2 + NAD+
show the reaction diagram
FMN + NADH
FMNH2 + NAD+
show the reaction diagram
FMN + NADH + H+
FMNH2 + NAD+
show the reaction diagram
lumiflavin + NADH + H+
reduced lumiflavin + NAD+
show the reaction diagram
-
6% of the activity with FAD
-
-
?
riboflavin + NADH + H+
reduced riboflavin + NAD+
show the reaction diagram
-
9% of the activity with FAD
-
-
?
additional information
?
-
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 + NAD(P)H
FADH2 + NAD(P)+
show the reaction diagram
-
-
-
-
?
FAD + NADH + H+
FADH2 + NAD+
show the reaction diagram
-
involved in pyrrolnitrin biosynthesis. PrnF is the flavin:NADH reductase component of the two-component arylamine oxygenase system in Pseudomonas fluorescens Pf-5. PrnF reduces FAD to FADH2, which is then directly transferred to PrnD, where it is used by PrnD to catalyze the oxygenation of aminopyrrolnitrin. The PrnD oxygenase component requires a direct interaction with the PrnF reductase component to oxygenate arylamine
-
-
?
additional information
?
-
-
the enzyme supplies reduced FADH2 for other enzymes, e.g. the 4-hydroxylphenylacetate 3-monooxygenase HpaB, which contains bound FADH2 and which protects FADH2 from being oxidized by O2, HpaC binds to HpaB without substrate channeling
-
-
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
FMN
-
12% of the activity with FAD
Lumiflavin
-
6% of the activity with FAD
NADPH
riboflavin
-
9% of the activity with FAD
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
AMP
-
AMP exhibits competitive inhibition versus NADH and noncompetitive inhibition versus FAD
HpaB
-
presence and binding of HpaB reduced the enzyme activity of HpaC due to stabilization and reduced oxidation by O2 of FADH2
-
NAD+
-
NAD+ demonstrates competitive inhibition versus NADH and noncompetitive inhibition versus FAD
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0008 - 0.056
FAD
0.0286
FMN
-
pH 7.0, 30C
0.0584
Lumiflavin
-
pH 7.0, 30C
0.008 - 0.183
NADH
0.0383
riboflavin
-
pH 7.0, 30C
additional information
additional information
-
kinetics, binding studies of HpaC and HpaB
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
40 - 178
FAD
72.4
FMN
Pseudomonas protegens Pf-5
-
pH 7.0, 30C
66.5
Lumiflavin
Pseudomonas protegens Pf-5
-
pH 7.0, 30C
40 - 178
NADH
68.2
riboflavin
Pseudomonas protegens Pf-5
-
pH 7.0, 30C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.318
-
strain W-KO, recombinant enzyme Fre, growth on 4-hydroxyphenylacetate as carbon source
25.1
-
strain W-KO, recombinant enzyme HpaC, growth on 4-hydroxyphenylacetate as carbon source
additional information
-
activity in a coupled assay of HpaB and HpaC
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.2
-
assay at
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 8
-
pH 6.0: 73% of maximal activity, pH 8.0: 81% of maximal activity
6.5 - 7.5
-
optimal range
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30
-
assay at, enzyme HpaC
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.3
-
calculated from sequence
6.5
-
isoelectric focusing
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
additional information
-
enzyme activities with different growth conditions, overview, strain W-KO does not grow on 4-hydroxyphenylacetate as carbon source
Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
18000
-
SDS-PAGE
19400
-
deduced from cDNA, without His-tag but with FAD
39000
-
gel filtration
additional information
-
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
dimer
-
2 * 19000, calculation from sequence
monomer
-
gel filtration
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
complexed with FMN, FAD or riboflavin, sitting drop vapor diffusion method, using 1.6 M ammonium sulfate and 4-5% (v/v) isopropanol
-
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4
-
pH 7.0, 1 mM dithiothreitol, 50 mM Tris-HCl, 50% loss of activity after 12 days
16
-
pH 7.0, 1 mM dithiothreitol, 50 mM Tris-HCl, 50% loss of activity after 5 days
30
-
pH 7.0, 1 mM dithiothreitol, 50 mM Tris-HCl, 50% loss of activity after 1 day
42
-
pH 7.0, 1 mM dithiothreitol, 50 mM Tris-HCl, 50% loss of activity after 30 min
50
-
pH 7.0, 1 mM dithiothreitol, 50 mM Tris-HCl, 50% loss of activity after 1 min
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Ni-affinity chromatography and heat treatment
-
recombinant His-tagged wild-type enzymes HpaC and Fre from strain BL21(DE3)
-
the reductase is purified as a stable C-terminally His-tagged yellow protein containing weakly bound FAD
-
using Ni-NTA chromatography
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Escherichia coli strain BL21(DE3)pLysS effectively produces an active and soluble form of StyB as about 9% of the total protein content, when cultivated at 20C with 0.5 mM IPTG
-
expressed in Escherichia coli as a His-tagged fusion protein
-
expressed in Escherichia coli BL21(DE3) cells
-
expressed in Escherichia coli Rosetta(DE3) cells
-
overexpression in Escherichia coli in a soluble form
-
recombinant overexpression of His-tagged wild-type enzymes HpaC and Fre in strain BL21(DE3), complementation of the inactivation mutant by transient expression of different gene hpaC variants, overview
-
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
expression is strongly induced in an NAP-1 (AP-1-like transcription factor) dependent manner
-
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
-
construction of an HpaC inactivation mutant strain W-KO
Renatured/COMMENTARY
ORGANISM
UNIPROT
LITERATURE