1.18.1.2: ferredoxin-NADP+ reductase
This is an abbreviated version!
For detailed information about ferredoxin-NADP+ reductase, go to the full flat file.
Word Map on EC 1.18.1.2
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1.18.1.2
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nitrate
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ferredoxins
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denitrification
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denitrify
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reductases
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heme
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fad
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spinach
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chloroplast
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copper
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ammonia
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nitrous
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flavin
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dissimilatory
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photosystems
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flavodoxins
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denitrificans
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alcaligenes
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paracoccus
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n2o
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anabaena
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copper-containing
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thylakoids
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anammox
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p450scc
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semiquinone
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diaphorase
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viologen
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azurins
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ferredoxin-dependent
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stutzeri
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plant-type
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achromobacter
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flavoenzymes
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electron-transfer
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ammonia-oxidizing
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plastocyanin
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dinitrogen
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photoreduction
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gogat
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pregnenolone
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nitrate-induced
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xylosoxidans
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photoreduced
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wolinella
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isoalloxazine
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nitrate-reducing
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biotechnology
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nitrosomonas
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deoxyhemoglobin
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agriculture
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single-electron
- 1.18.1.2
- nitrate
- ferredoxins
-
denitrification
-
denitrify
- reductases
- heme
- fad
- spinach
- chloroplast
- copper
- ammonia
-
nitrous
- flavin
-
dissimilatory
- photosystems
- flavodoxins
- denitrificans
- alcaligenes
- paracoccus
- n2o
- anabaena
-
copper-containing
- thylakoids
-
anammox
- p450scc
- semiquinone
- diaphorase
- viologen
- azurins
-
ferredoxin-dependent
- stutzeri
-
plant-type
- achromobacter
-
flavoenzymes
-
electron-transfer
-
ammonia-oxidizing
- plastocyanin
-
dinitrogen
-
photoreduction
- gogat
- pregnenolone
-
nitrate-induced
- xylosoxidans
-
photoreduced
-
wolinella
- isoalloxazine
-
nitrate-reducing
- biotechnology
- nitrosomonas
-
deoxyhemoglobin
- agriculture
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single-electron
Reaction
2 reduced ferredoxin
+
Synonyms
ABO_0145, adrenodoxin reductase, AnFNR, BsFNR, CT1512, cytochrome b6f -associated ferredoxin:NADP+ oxidoreductase, DA1, EC 1.6.7.1, EC 1.6.99.4, Fd-NADP+ reductase, ferredoxin (flavodoxin)-(reduced) nicotinamide adenine dinucleotide phosphate reductase, ferredoxin (flavodoxin)-NAD(P)H reductase, ferredoxin (flavodoxin)-NADP(H) reductase, ferredoxin (flavodoxin):NADP+ oxidoreductase, ferredoxin NADP oxidoreductase, ferredoxin NADP reductase, ferredoxin NADP(H) oxidoreductase, ferredoxin NADP+ oxidoreductase, ferredoxin NADP+ reductase, ferredoxin nicotinamide adenine dinucleotide phosphate reductase, ferredoxin-NAD(P)+ oxidoreductase, ferredoxin-NAD(P)+ reductase, ferredoxin-NAD(P)H reductase, ferredoxin-NADP oxidoreductase, ferredoxin-NADP reductase, Ferredoxin-NADP(+) reductase, ferredoxin-NADP(H) oxidoreductase, ferredoxin-NADP(H) reductase, ferredoxin-NADP+ oxidoreductase, ferredoxin-NADP+ reductase, ferredoxin-NADP+-oxidoreductase, ferredoxin-NADP+-reductase, ferredoxin-NADP-oxidoreductase, ferredoxin-NADP-reductase, ferredoxin-nicotinamide adenine dinucleotide phosphate reductase, ferredoxin-nicotinamide-adenine dinucleotide phosphate (oxidized) reductase, ferredoxin-TPN reductase, ferredoxin: NADP(+) oxidoreductase, ferredoxin: NADP+ oxidoreductase, ferredoxin:NADP oxidoreductase, ferredoxin:NADP(+) oxidoreductase, ferredoxin:NADP(H) oxidoreductase, ferredoxin:NADP(H) oxidoreductase 2, ferredoxin:NADP+ oxidoreductase, ferredoxin:NADP+ reductase, ferredoxin:NADPH oxidoreductase, ferric reductase, Flavodoxin reductase, FLDR, FLXR, FNR, FNR-A, FNR-B, FNR1, FNR2, FNRII, FNRL, FNRS, FPR, FprA, FprB, Fprs, LFNR1, LFNR2, mitochondrial-type ferredoxin:NADP+ reductase, More, mtFNR, NADH-dependent reduced ferredoxin:NADP oxidoreductase, NADP:ferredoxin oxidoreductase, NADPH ferredoxin reductase, NADPH-dependent ferredoxin reductase, NADPH:ferredoxin oxidoreductase, NfnAB, NfnIII, NFR, PETH, PfFNR, pFNR, photosynthetic ferredoxin NADP+ oxidoreductase, photosynthetic FNR, plastidic-type ferredoxin-NADP+ reductase, PP4_41290, RcFPR, reduced nicotinamide adenine dinucleotide phosphate-adrenodoxin reductase, reductase, ferredoxin-nicotinamide adenine dinucleotide phosphate, RFNR2, ST2133, TgFNR, TPNH-ferredoxin reductase, YumC
ECTree
1.18.1.2 ferredoxin-NADP+ reductaseAdvanced search results
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results (Engineering
Engineering on EC 1.18.1.2 - ferredoxin-NADP+ reductase
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PROTEIN VARIANTS 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
E139D
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site-directed mutagenesis, altered conformation compared to the wild-type enzyme, slightly reduced activity compared to the wild-type enzyme
E139K
E139Q
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site-directed mutagenesis, mutant enzyme shows increased interaction with ferredoxin and reduces the appropriate orientation of flavodoxin, altered conformation compared to the wild-type enzyme, increased activity compared to the wild-type enzyme
E301A
I59A
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kcat for flavodoxin is slightly decreased, but kcat/Km is markedly increased
K138E
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site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
K290E
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site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
K294E
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site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
K72E
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site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
K75E
L263A
site-directed mutagenesis, altered cofactor specificity compared to the wild-type enzyme
L263P
site-directed mutagenesis, altered cofactor specificity compared to the wild-type enzyme
L76D
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site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
L76F
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site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
L76S
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site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
L76V
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site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
L78D
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site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
L78F
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site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
L78S
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site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
L78V
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site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
R100A
R16E
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site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
R224Q/R233L/Y235F
site-directed mutagenesis, altered cofactor specificity compared to the wild-type enzyme
R233L/Y235F
site-directed mutagenesis, altered cofactor specificity compared to the wild-type enzyme
R264E
S223D/R224Q/R233L/Y235F
site-directed mutagenesis, altered cofactor specificity compared to the wild-type enzyme
S223D/R233L/Y235F
site-directed mutagenesis, altered cofactor specificity compared to the wild-type enzyme
T155G
site-directed mutagenesis, altered cofactor specificity compared to the wild-type enzyme
T155G/A160T
site-directed mutagenesis, altered cofactor specificity compared to the wild-type enzyme
T155G/A160T/L263P
T155G/A160T/L263P/R264P/G265P
called PP5, increased Km but reduced kcat for NADPH
T155G/A160T/L263P/Y303S
called PP3CT, catalytic efficiency comparable with wild type, high specificity for NADH
T155G/A160T/S223D/R224Q/R233L/Y235F
site-directed mutagenesis, altered cofactor specificity compared to the wild-type enzyme
T155G/A160T/S223D/R224Q/R233L/Y235F/L263P
site-directed mutagenesis, altered cofactor specificity compared to the wild-type enzyme
T155G/A160T/S223D/Y235F/L263P/R264P/G265P
called AMP2PP5, almost no reactivity
T155G/R224Q/R233L/Y235F
site-directed mutagenesis, altered cofactor specificity compared to the wild-type enzyme
T155G/S223D/R224Q/R233L/Y235F
site-directed mutagenesis, altered cofactor specificity compared to the wild-type enzyme
Y303F
Y303S
Y303W
E301A
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8% FAD semiquinone at the equilibrium. Mutation does not change quinone substrate specificity but confers the mixed single- and two-electron mechanism of quinone reduction, whereas wild-type uses a single-electron pathway. Change can be explained by the relative increase in the rate of second electron transfer
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H324F
site-directed mutagenesis, the mutant shows altered kinetics and reduced activity compared to the wild-type FNR
H324S
site-directed mutagenesis, the mutant shows altered kinetics and reduced activity compared to the wild-type FNR
R186G
site-directed mutagenesis, replacement of Arg186 with glycine leads to drastically reduced amounts of recombinant protein
R186G/D187H
site-directed mutagenesis, the mutant shows altered kinetics and reduced activity compared to the wild-type FNR
R186G/D187H/R190Q
site-directed mutagenesis, the mutant shows altered kinetics and reduced activity compared to the wild-type FNR
R190Q
site-directed mutagenesis, the mutant shows altered kinetics and reduced activity compared to the wild-type FNR
Y50G
Y50S
Y50W
Y50G
Y50S
Y50W
K135L
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site-directed mutagenesis, altered tertiary structure compared to the wild-type enzyme, no methylation at K83, 10% of native wild-type enzyme activity
K83L
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site-directed mutagenesis, altered tertiary structure compared to the wild-type enzyme, no methylation at K83, 2-4% of native wild-type enzyme activity
K83L/K89L
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site-directed mutagenesis, altered tertiary structure compared to the wild-type enzyme, no methylation at K83, 0.3% of native wild-type enzyme activity
K89L
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site-directed mutagenesis, altered tertiary structure compared to the wild-type enzyme, no methylation at K83, 4% of native wild-type enzyme activity
Y308S
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
Y308F
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site-directed mutagenesis, about 20% of the wild-type enzyme activity with ferredoxin, about 11% of the wild-type enzyme activity with flavodoxin
Y308S
Y308W
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site-directed mutagenesis, nearly inactive mutant with ferredoxin, about 25% of the wild-type enzyme activity with flavodoxin
H286K
considerable decrease in kcat and kcat/Km, almost no reactivity with K3Fe(CN)6
H286L
kcat and kcat/Km are markedly reduced, anomalous reactivity with 2,6-dichlorophenolindophenol
H286Q
up to 50% higher kcat than wild type, higher kcat/Km when 2,6-dichlorophenolindophenol serves as electron acceptor
K259A
50% decrease in catalytic efficiency compared to wild-type
K259D
50% decrease in catalytic efficiency compared to wild-type
A266Y
A266y/Del267-272
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
deletion emulates the structure present in plastidic versions of the protein, mutant displays higher affinity for NADP+ than wild-type
DELTA1-15
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removal of the N-terminal 15 residues of enzyme enhances the interaction with ferredoxin
DELTA81-118
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deleted amino acid comprise a species-specific subdomain containing an insertion of 28 residues, deletion results in an catalytically active enzyme form with 10fold decreased affinity for ferredoxin
Q242R
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catalytic activity similar to wild-type. Protein moves faster than wild-type on SDS-PAGE
Q242R/S267R
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10fold increase in binding affinity for ferredoxin, 1-4% of wild-type activity.
S267R
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10fold increase in binding affinity for ferredoxin, 1-4% of wild-type activity
E19C
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site-directed mutagenesis of isozyme L-FNR I, NADPH-dependent cyt c reduction activity with different recombinant wild-type and mutant ferredoxins, in comparison to the wild-type enzyme, NMR chemical shift perturbation analysis, overview
E25C
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site-directed mutagenesis of isozyme L-FNR I, NADPH-dependent cyt c reduction activity with different recombinant wild-type and mutant ferredoxins, in comparison to the wild-type enzyme, NMR chemical shift perturbation analysis, overview
E36C
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site-directed mutagenesis of isozyme L-FNR I, NADPH-dependent cyt c reduction activity with different recombinant wild-type and mutant ferredoxins, in comparison to the wild-type enzyme, NMR chemical shift perturbation analysis, overview
additional information
E139K
-
site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
E139K
-
site-directed mutagenesis, mutant enzyme shows increased interaction with ferredoxin and reduces the appropriate orientation of flavodoxin, altered conformation compared to the wild-type enzyme, increased activity compared to the wild-type enzyme
E301A
-
site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
E301A
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8% FAD semiquinone at the equilibrium. Mutation does not change quinone substrate specificity but confers the mixed single- and two-electron mechanism of quinone reduction, whereas wild-type uses a single-electron pathway. Change can be explained by the relative increase in the rate of second electron transfer
E301A
-
site-directed mutagensis, kinetic parameters for hydride and deuteride transfer processes between enzyme and NADP+ in comparison to the wild-type enzyme, overview
K75E
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site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
K75E
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mutation results in changes of charge distribution on the surface of the protein and in formation of a salt bridge between E75 and K72 that inhibits the essential interaction of these residues with flavodoxin/ferredoxin
R100A
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amino acid replacement removes the positive charge and the ability to form hydrogen bonds, enzyme interacts weakly with Cibacron-Blue Sepharose, increased Km for NADPH
R100A
-
site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
R100A
-
site-directed mutagensis, kinetic parameters for hydride and deuteride transfer processes between enzyme and NADP+ in comparison to the wild-type enzyme, overview
R264E
-
site-directed mutagenesis, altered interaction and kinetics of enzyme-ferredoxin association compared to the wild-type enzyme
T155G/A160T/L263P
site-directed mutagenesis, altered cofactor specificity compared to the wild-type enzyme
T155G/A160T/L263P
called PP3, only half catalytic efficiency compared with wild type
Y303F
site-directed mutagenesis, about 30% of the wild-type enzyme activity with ferredoxin, about 25% of the wild-type enzyme activity with flavodoxin
Y303F
-
site-directed mutagensis, kinetic parameters for hydride and deuteride transfer processes between enzyme and NADP+ in comparison to the wild-type enzyme, overview
Y303S
-
site-directed mutagensis, kinetic parameters for hydride and deuteride transfer processes between enzyme and NADP+ in comparison to the wild-type enzyme, overview
Y303W
-
site-directed mutagensis, kinetic parameters for hydride and deuteride transfer processes between enzyme and NADP+ in comparison to the wild-type enzyme, overview
Y50G
mutation results in a blue shift of the FAD transition bands, with enhancement of fluorescence emission. Mutant displays decreased thermal stability
Y50G
site-directed mutagenesis, the mutant shows a blue shift of the FAD transition band and decreased thermal stability compared to wild-type. Using the diaphorase assay, the kcat values for the Y50G mutant in the presence of NADPH and ferricyanide is decreased to less than 5% of the wild-type activity
Y50G
the mutation decreases thermal stability compared to the wild type enzyme
Y50S
mutation results in a blue shift of the FAD transition bands, with enhancement of fluorescence emission. Mutant displays decreased thermal stability
Y50S
site-directed mutagenesis, the mutant shows a blue shift of the FAD transition band and decreased thermal stability compared to wild-type. Using the diaphorase assay, the kcat values for the Y50G mutant in the presence of NADPH and ferricyanide is decreased to less than 5% of the wild-type activity
Y50S
the mutation decreases thermal stability compared to the wild type enzyme
Y50W
mutation results in a blue shift of the FAD transition bands, with quenching of fluorescence emission. Mutant displays decreased thermal stability
Y50W
site-directed mutagenesis, the mutant shows a blue shift of the FAD transition band and decreased thermal stability compared to wild-type. The mutant retains approximately 20 % reactivity in the diaphorase assay and BsFd-dependent cytochrome c reduction assay relative to wild-type
Y50W
the mutation decreases thermal stability compared to the wild type enzyme, The mutant retains approximately 20% of wild type reactivity
Y50G
-
site-directed mutagenesis, the mutant shows a blue shift of the FAD transition band and decreased thermal stability compared to wild-type. Using the diaphorase assay, the kcat values for the Y50G mutant in the presence of NADPH and ferricyanide is decreased to less than 5% of the wild-type activity
-
Y50G
-
mutation results in a blue shift of the FAD transition bands, with enhancement of fluorescence emission. Mutant displays decreased thermal stability
-
Y50S
-
site-directed mutagenesis, the mutant shows a blue shift of the FAD transition band and decreased thermal stability compared to wild-type. Using the diaphorase assay, the kcat values for the Y50G mutant in the presence of NADPH and ferricyanide is decreased to less than 5% of the wild-type activity
-
Y50S
-
mutation results in a blue shift of the FAD transition bands, with enhancement of fluorescence emission. Mutant displays decreased thermal stability
-
Y50W
-
site-directed mutagenesis, the mutant shows a blue shift of the FAD transition band and decreased thermal stability compared to wild-type. The mutant retains approximately 20 % reactivity in the diaphorase assay and BsFd-dependent cytochrome c reduction assay relative to wild-type
-
Y50W
-
mutation results in a blue shift of the FAD transition bands, with quenching of fluorescence emission. Mutant displays decreased thermal stability
-
Y308S
-
altered cofactor specificity compared to the wild-type enzyme, mutant enzymes is able to utilizes NADP(H) as well as NAD(H)
Y308S
-
site-directed mutagenesis, about 5% of the wild-type enzyme activity with ferredoxin, no activity with flavodoxin
A266Y
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
site-directed mutagenesis, deletion of the six C-terminal amino acids FVGEGI beyond Ala266 is combined with the replacement A266Y to emulate the structure of plastidic reductases. THe mutations produce subtle global conformational changes, but strongly reduce the local rigidity of the FAD-binding pocket, exposing the isoalloxazine ring to the solvent. Thus, the ultrafast charge-transfer quenching of FAD* by the conserved Tyr66 residue is absent in the mutant series, producing enhancement of the excited singlet and triplet-state properties of FAD. All RcFPR variants display higher affinity for NADP+ than the wild-type
additional information
isozyme LFNR2 can only partially compensate for the function of LFNR1
additional information
isozyme LFNR2 can only partially compensate for the function of LFNR1
additional information
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isozyme LFNR2 can only partially compensate for the function of LFNR1
additional information
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construction of in Arabidopsis mutants completely devoid of FNR1 or FNR2
additional information
site-directed mutagenesis of NADPH-specific BsFNR to replace Arg186, Asp187, Arg190, and His324 with the residues occurring in NADH/NADPH-bispecific Chlorobium tepidum FNR
additional information
-
site-directed mutagenesis of NADPH-specific BsFNR to replace Arg186, Asp187, Arg190, and His324 with the residues occurring in NADH/NADPH-bispecific Chlorobium tepidum FNR
additional information
-
replacement of Tyr50 stacked on the si-face of the isoalloxazine ring of the flavin adenine dinucleotide prosthetic group modulates Bacillus subtilis ferredoxin-NADP+ oxidoreductase activity toward NADPH. The Y50G and Y50S mutations enhance the FAD fluorescence emission, whereas those of the wild type and Y50W mutant are quenched
additional information
replacement of Tyr50 stacked on the si-face of the isoalloxazine ring of the flavin adenine dinucleotide prosthetic group modulates Bacillus subtilis ferredoxin-NADP+ oxidoreductase activity toward NADPH. The Y50G and Y50S mutations enhance the FAD fluorescence emission, whereas those of the wild type and Y50W mutant are quenched
additional information
-
the depletions of residues Y313 to K332 (whole C-terminal extension region) and S325 to K332 results in significant increases in the catalytic efficiency with NADPH in diaphorase assay with ferricyanide, whereas Km values for ferricyanide are increased. In the cytochrome c reduction assay in the presence of ferredoxin, the S325-K332 depleted mutant displays a significant decrease in the turnover rate. The Y313-K332 depleted mutant demonstrates an increase in the rate of the direct reduction of horse heart cytochrome c in the absence of ferredoxin
additional information
-
the depletions of residues Y313 to K332 (whole C-terminal extension region) and S325 to K332 results in significant increases in the catalytic efficiency with NADPH in diaphorase assay with ferricyanide, whereas Km values for ferricyanide are increased. In the cytochrome c reduction assay in the presence of ferredoxin, the S325-K332 depleted mutant displays a significant decrease in the turnover rate. The Y313-K332 depleted mutant demonstrates an increase in the rate of the direct reduction of horse heart cytochrome c in the absence of ferredoxin
-
additional information
-
replacement of Tyr50 stacked on the si-face of the isoalloxazine ring of the flavin adenine dinucleotide prosthetic group modulates Bacillus subtilis ferredoxin-NADP+ oxidoreductase activity toward NADPH. The Y50G and Y50S mutations enhance the FAD fluorescence emission, whereas those of the wild type and Y50W mutant are quenched
-
additional information
-
deletions of Tyr326-Glu360 decrease the hydride transfer rate and the amount of reduced enzyme increases at equilibrium relative to wild type. Deletions of Phe337-Glu360 and Ser338-Glu360 result in slight changes in the reaction kinetics and redox equilibrium compared to the wild type enzyme
additional information
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construction of ferredoxin II mutants Q39R/S28E and D64N by site-directed mutagenesis, expression of recombinant wild-type and mutant ferredoxins of Equisetum arvense in Escherichia coli
additional information
mutual exchange of the 112-123 beta-hairpin from Pisum sativum plastidic ferredoxinNAD(P)H reductase and the carboxy-terminal tryptophan of he Escherichia coli enzyme. The plastidic enzyme lacking the beta-hairpin is unable to fold properly. An extra tryptophan at the carboxy terminus, emulating the bacterial enzyme, results in an enzyme with decreased affinity for FAD and reduced diaphorase and ferredoxin-dependent cytochrome c reductase activities. The insertion of the beta-hairpin into the corresponding position of the bacterial enzyme increases FAD affinity but does not affect its catalytic properties. The same insertion with simultaneous deletion of the carboxyterminal tryptophan produces a bacterial chimera emulating the plastidic architecture with an increased kcat and an increased catalytic efficiency for the diaphorase activity and a decrease in the enzymes ability to react with its substrates ferredoxin and flavodoxin. Crystallographic structures of the chimeras show no significant changes in their overall structure, although alterations in the FAD conformations are observed
additional information
-
addition of an artificial metal binding site of nine amino acids, including four His residues, to the C-terminal Tyr308 residue. The additional structure binds Zn2+ or Co2+ and significantly reduces the catalytic efficiency of the enzyme by decreasing the kcat value. In absence of Zn2+, Km value of NADPH and Kd value for NADP+ are increased 2 to 3 times
additional information
mutual exchange of the 112-123 beta-hairpin from Pisum sativum plastidic ferredoxinNAD(P)H reductase and the carboxy-terminal tryptophan of he Escherichia coli enzyme. The plastidic enzyme lacking the beta-hairpin is unable to fold properly. An extra tryptophan at the carboxy terminus, emulating the bacterial enzyme, results in an enzyme with decreased affinity for FAD and reduced diaphorase and ferredoxin-dependent cytochrome c reductase activities. The insertion of the beta-hairpin into the corresponding position of the bacterial enzyme increases FAD affinity but does not affect its catalytic properties. The same insertion with simultaneous deletion of the carboxyterminal tryptophan produces a bacterial chimera emulating the plastidic architecture with an increased kcat and an increased catalytic efficiency for the diaphorase activity and a decrease in the enzyme's ability to react with its substrates ferredoxin and flavodoxin. Crystallographic structures of the chimeras show no significant changes in their overall structure, although alterations in the FAD conformations are observed
additional information
-
a tyrosine mutant accepts NAD(H) as cofactor and is insensitive to inhibition by NADH
additional information
-
construction of 2 different chimeric enzymes with domains of spinach nonphotosynthetic root isozyme and photosynthetic leaf isozyme, the chimera comprising the root NADP-binding domain and the leaf FAD-binding domain is functional, while the chimera with opposite composition is not, the active chimera shows partially impaired intermolecular electron transfer, and highly increased NADPH-diaphorase activity, and increased activity with NADH as cofactor compared to the parent isozymes, improvement of catalytic properties, overview
additional information
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study on impact of N-terminal truncation on interaction with ferredoxin using deletions of up to 25 amino acids. Similar kcat values for cytochrome c reduction are measured for all but the most truncated pFNRII[N-5]DEGV which contains a truncated N-terminal domain beginning after the KISKK domain. The longer forms of isoform FNRII bind more strongly to a ferredoxin affinity column than the shorter forms. Mutants bearing lysine-to-glutamine mutations in the KISKK domain are affected in binding to the ferredoxin column
