Information on EC 1.19.1.1 - flavodoxin-NADP+ reductase

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

EC NUMBER
COMMENTARY hide
1.19.1.1
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RECOMMENDED NAME
GeneOntology No.
flavodoxin-NADP+ reductase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
reduced flavodoxin + NADP+ = oxidized flavodoxin + NADPH + H+
show the reaction diagram
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SYSTEMATIC NAME
IUBMB Comments
flavodoxin:NADP+ oxidoreductase
A flavoprotein (FAD). This activity occurs in some prokaryotes and algae that possess flavodoxin, and provides low-potential electrons for a variety of reactions such as nitrogen fixation, sulfur assimilation and amino acid biosynthesis. In photosynthetic organisms it is involved in the photosynthetic electron transport chain. The enzyme also catalyses EC 1.18.1.2, ferredoxin---NADP+ reductase.
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
enzyme catalyzes reactions of both EC 1.18.1.2 and EC 1.19.1.1
UniProt
Manually annotated by BRENDA team
enzyme catalyzes reactions of both EC 1.18.1.2 and EC 1.19.1.1
UniProt
Manually annotated by BRENDA team
enzyme catalyzes reactions of both EC 1.18.1.2 and EC 1.19.1.1
UniProt
Manually annotated by BRENDA team
enzyme catalyzes reactions of both EC 1.18.1.2 and EC 1.19.1.1
UniProt
Manually annotated by BRENDA team
enzyme catalyzes reactions of both EC 1.18.1.2 and EC 1.19.1.1
UniProt
Manually annotated by BRENDA team
enzyme catalyzes reactions of both EC 1.18.1.2 and EC 1.19.1.1
Uniprot
Manually annotated by BRENDA team
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 ferricyanide + NADPH
2 ferrocyanide + NADP+ + H+
show the reaction diagram
2 ferricyanide + NADPH + H+
2 ferrocyanide + NADP+ + H+
show the reaction diagram
2 ferricytochrome c + NADPH + H+
2 ferrocytochrome c + NADP+ + H+
show the reaction diagram
2 ferricytochrome c2 + NADPH
2 ferrocytochrome c2 + NADP+ + H+
show the reaction diagram
2,6-dichlorophenolindophenol + NADPH
reduced 2,6-dichlorophenolindophenol + NADP+
show the reaction diagram
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-
-
-
?
FMNH2 + NADP+
FMN + NADPH + H+
show the reaction diagram
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-
-
?
oxidized cytochrome c + NADH + H+
reduced cytochrome c + NAD+
show the reaction diagram
oxidized cytochrome c + NADPH + H+
reduced cytochrome c + NADP+
show the reaction diagram
reduced 2,6-dichlorophenolindophenol + NADP+
oxidized 2,6-dichlorophenolindophenol + NADPH + H+
show the reaction diagram
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-
-
-
?
reduced flavodoxin + NADP+
oxidized flavodoxin + NADPH + H+
show the reaction diagram
reduced flavodoxin I + NADP+
oxidized flavodoxin I + NADPH + H+
show the reaction diagram
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?
reduced flavodoxin II + NADP+
oxidized flavodoxin II + NADPH + H+
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
FMNH2 + NADP+
FMN + NADPH + H+
show the reaction diagram
P28861
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-
-
?
reduced flavodoxin I + NADP+
oxidized flavodoxin I + NADPH + H+
show the reaction diagram
P28861
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?
reduced flavodoxin II + NADP+
oxidized flavodoxin II + NADPH + H+
show the reaction diagram
P28861
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?
additional information
?
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P28861
the enzyme reduces flavodoxin I, flavodoxin II and ferredoxin, ferredoxin being the kinetically and thermodynamically preferred partner, i.e. reaction of EC 1.18.2.1. Flavodoxin I and flavodoxin II behave similarly with respect to FNR, with affinities about 4- to 7fold weaker and reduction rates that are 10- to 100fold slower than those for ferredoxin. Flavodoxin I and flavodoxin II can obtain electrons from reduced Fd at rates that are comparable to those obtained with reduced FNR
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
NADP+
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
flavodoxin
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cytochrome c reductase activity of FPR is stimulated three times by the addition of flavodoxin
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0236
ferricyanide
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pH 7.5, 30C
0.009
ferricytochrome c
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presence of flavodoxin, pH 8.0, temperature not specified in the publication
0.0176
ferricytochrome c2
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pH 7.5, 30C
0.0017 - 0.0099
NADH
0.0039 - 0.093
NADPH
0.0068 - 0.043
reduced flavodoxin
0.0076
reduced flavodoxin I
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pH 8.0, 25C
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0.004
reduced flavodoxin II
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pH 8.0, 25C
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
26.8
ferricyanide
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pH 7.5, 30C
0.25
ferricytochrome c
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presence of flavodoxin, pH 8.0, temperature not specified in the publication
2.35
ferricytochrome c2
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pH 7.5, 30C
0.23 - 0.84
NADH
1 - 222
NADPH
2.5 - 30.6
reduced flavodoxin
0.0042
reduced flavodoxin I
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pH 8.0, 25C
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0.0039
reduced flavodoxin II
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pH 8.0, 25C
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kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
19000
FMNH2
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pH 8.0, 25C
270 - 715
NADH
93 - 1448
NADPH
170 - 1800
reduced flavodoxin
550
reduced flavodoxin I
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pH 8.0, 25C
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980
reduced flavodoxin II
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pH 8.0, 25C
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pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4.8
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isoelectric focusing
6.2
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calculated
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30000
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gel filtration
80000 - 110000
SDS-PAGE
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
additional information
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
structure to 2.5 A resolution, orthorhombic space group P21212, with unit-cell parameters a = 57.2, b = 164.3, c = 95.0 A, containing two protein molecules in the asymmetric unit
molecular modelling indicates that movement of the C-terminal tryptophan (W248) is necessary to permit close approach of the nicotinamide ring of NADPH to the flavin. Residues R174 and R184 are located close to the adenosine ribose 2'-phosphate group, and R144 is likely to interact with the nicotinamide ribose 5'-phosphate group
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a multiscale modelling approach for analysis of the electron transfer process in complexes of the enzyme with both ferredoxin and flavodoxin, reactions of EC 1.19.1.1 and EC1.18.1.2, respectively. The electron transfer in FNR/ferredoxin proceedes through a bridge-mediated mechanism in a dominant protein-protein complex, where transfer of the electron is facilitated by ferredoxin loop-residues 40-49. In FNR/flavodoxin, a direct transfer between redox cofactors is observed and less complex specificity than in ferredoxin
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in complex with 2'-phospho-AMP and NADP+. In the complexes obtained, the nucleotides bind exclusively through the adenosine moiety. The adenosine moiety binds into a cavity formed by residues of conserved segments, i.e residues 128 to 130, residues 158 to 163, residues 193 to 205, and residues 233 to 240. The adenosine binding site is essentially formed by residues R158, R195 and R203, which stabilise the nucleotide
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to 2.17 A resolution, tetragonal space group P41212, with unit-cell parameters a = b = 66.49, c = 121.32 A
no perturbation of the 31P-NMR resonances assigned to the FAD moiety of FNR or the FMN and phosphodiester moieties of Azotobacter flavodoxin are observed on complexation of Azotobacter flavodoxin and Spinacia oleracea FNR. Reduction of FMN to its semiquinone form results in extensive line-broadening of the FMN resonance. The FAD resonances of the FNR-flavodoxin complex are unaffected by FMN semiquinone formation. The distance from the FMN phosphate to the flavin ring is altered on binding the flavodoxin to FNR
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37
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reduced FNR is subject to inactivation due to unfolding of the protein and dissociation of the FADH2 cofactor
41
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melting temperature,reduced FNR in presence of dithiothreitol
66
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melting temperature,oxidized FNR in presence of dithiothreitol
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
binding of ferredoxin, FAD, flavodoxin, or riboflavin stabilizes the enzyme
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FNR reduced in the presence of NADPH is slowly inactivated under all conditions. Reactivity towards flavodoxin is lost most rapidly (kinact of 0.031 per min) with less than 10% of the original activity remaining after 30 min, reactivity towards ferredoxin is not as rapidly affected (kinact of 0.0065 per min) with 80% of the original activity remaining after 30 min
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
recpmbinant protein. FLDR is bright yellow in its oxidized form and it is converted to a neutral blue semiquinone by the addition of one reducing equivalent
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Y50G
mutation results in a blue shift of the FAD transition bands, with enhancement of fluorescence emission. Mutant displays decreased thermal stability
Y50S
mutation results in a blue shift of the FAD transition bands, with enhancement of fluorescence emission. Mutant displays decreased thermal stability
Y50W
mutation results in a blue shift of the FAD transition bands, with quenching of fluorescence emission. Mutant displays decreased thermal stability
Y50G
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mutation results in a blue shift of the FAD transition bands, with enhancement of fluorescence emission. Mutant displays decreased thermal stability
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Y50S
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mutation results in a blue shift of the FAD transition bands, with enhancement of fluorescence emission. Mutant displays decreased thermal stability
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R144A
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mutation in the proposed NADPH-binding site, mutant exhibits decreased NADPH-dependent cytochrome c reductase activity and increased Km for NADPH
R174A
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mutation in the proposed NADPH-binding site, mutant exhibits decreased NADPH-dependent cytochrome c reductase activity and increased Km for NADPH
R184A
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mutation in the proposed NADPH-binding site, mutant exhibits decreased NADPH-dependent cytochrome c reductase activity and increased Km for NADPH
Y308S
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mutant uses NAD(H) instead of NADP(H), expression of the mutant has no effect on soxRS induction and fails to protect FPR deficient cells from methyl viologen toxicity
R144A
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mutation in the proposed NADPH-binding site, mutant exhibits decreased NADPH-dependent cytochrome c reductase activity and increased Km for NADPH
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R174A
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mutation in the proposed NADPH-binding site, mutant exhibits decreased NADPH-dependent cytochrome c reductase activity and increased Km for NADPH
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R184A
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mutation in the proposed NADPH-binding site, mutant exhibits decreased NADPH-dependent cytochrome c reductase activity and increased Km for NADPH
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Y303F
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about 30% of the wild-type enzyme activity with ferredoxin, about 25% of the wild-type enzyme activity with flavodoxin
Y303S
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inactive. Mutation shifts the flavin reduction potential to less negative values, whereas semiquinone stabilization is severely hampered
Y303W
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almost inactive
Y303F
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about 30% of the wild-type enzyme activity with ferredoxin, about 25% of the wild-type enzyme activity with flavodoxin
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Y303S
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inactive. Mutation shifts the flavin reduction potential to less negative values, whereas semiquinone stabilization is severely hampered
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Y303W
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almost inactive
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Y308F
about 20% of the wild-type enzyme activity with ferredoxin, about 11% of the wild-type enzyme activity with flavodoxin
Y308S
nearly inactive mutant with ferredoxin, about 25% of the wild-type enzyme activity with flavodoxin. Mutation shifts the flavin reduction potential to less negative values, whereas semiquinone stabilization is severely hampered
Y308W
about 5% of the wild-type enzyme activity with ferredoxin, no activity with flavodoxin
A266Y
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mutant does not allow formation of active charge-transfer complexes, probably due to restraints of C-terminus pliability. Mutant displays higher affinity for NADP+ than wild-type
A266y/Del267-272
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deletion/mutation emulates the structure present in plastidic versions of the protein. It does not modify the general geometry of FAD itself, but increases exposure of the flavin to the solvent, prevents a productive geometry of FAD:NADP(H) complex and decreases the protein thermal stability. Mutant displays higher affinity for NADP+ than wild-type
Del267-272
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deletion emulates the structure present in plastidic versions of the protein, mutant displays higher affinity for NADP+ than wild-type