Information on EC 1.8.1.2 - assimilatory sulfite reductase (NADPH)

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

EC NUMBER
COMMENTARY hide
1.8.1.2
-
RECOMMENDED NAME
GeneOntology No.
assimilatory sulfite reductase (NADPH)
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
hydrogen sulfide + 3 NADP+ + 3 H2O = sulfite + 3 NADPH + 3 H+
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
oxidation
redox reaction
-
-
-
-
reduction
additional information
-
sodium salts of thiosulfate and sulfate do not serve as the electron acceptor for reduced F420 oxidation by Fsr. Also, Fsr can not use NADH and NADPH for the reduction of sulfite.; the N-terminal half of Fsr represents a H2F420 dehydrogenase and the C-terminal half a dissimilatory-type siroheme sulfite reductase, and Fsr catalyzes the corresponding partial reactions
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Metabolic pathways
-
-
Microbial metabolism in diverse environments
-
-
sulfate reduction
-
-
sulfate reduction I (assimilatory)
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sulfate reduction III (assimilatory)
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-
Sulfur metabolism
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-
SYSTEMATIC NAME
IUBMB Comments
hydrogen-sulfide:NADP+ oxidoreductase
Contains siroheme, [4Fe-4S] cluster, FAD and FMN. The enzyme, which catalyses the six-electron reduction of sulfite to sulfide, is involved in sulfate assimilation in bacteria and yeast. Different from EC 1.8.99.5, dissimilatory sulfite reductase, which is involved in prokaryotic sulfur-based energy metabolism. cf. EC 1.8.7.1, assimilatory sulfite reductase (ferredoxin).
CAS REGISTRY NUMBER
COMMENTARY hide
9029-35-0
-
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
1,1'-dimethyl-4,4'-bipyridinium dichloride + NADPH
NADPH + oxidized 1,1'-dimethyl-4,4'-bipyridinium dichloride
show the reaction diagram
-
i.e. paraquat, a herbicide
-
-
-
hydrogen sulfide + 3 NADP+ + 3 H2O
sulfite + 3 NADPH + 3 H+
show the reaction diagram
-
-
-
-
?
hydrogen sulfide + NADP+ + H2O
sulfite + NADPH + H+
show the reaction diagram
-
-
-
-
?
hydroxylamine + NADPH
ammonia + NADP+
show the reaction diagram
NADP+ + reduced methyl viologen
NADPH + oxidized methyl viologen
show the reaction diagram
nitrite + NADPH
ammonia + NADP+
show the reaction diagram
sulfite + NADPH
hydrogen sulfide + NADP+ + H2O
show the reaction diagram
-
-
-
-
?
sulfite + NADPH
sulfide + NADP+ + H2O
show the reaction diagram
sulfite + NADPH + H+
sulfide + NADP+ + H2O
show the reaction diagram
sulfite + reduced F420
sulfide + oxidized F420
show the reaction diagram
-
-
-
-
?
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
hydrogen sulfide + 3 NADP+ + 3 H2O
sulfite + 3 NADPH + 3 H+
show the reaction diagram
-
-
-
-
?
hydrogen sulfide + NADP+ + H2O
sulfite + NADPH + H+
show the reaction diagram
-
-
-
-
?
sulfite + NADPH
sulfide + NADP+ + H2O
show the reaction diagram
sulfite + NADPH + H+
sulfide + NADP+ + H2O
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
flavin
iron-sulfur centre
siroheme
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Iron-sulfur cluster
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2'-AMP
-
NADPH-sulfite reductase and NADPH-cytochrome c reductase
8-hydroxyquinoline
-
-
arsenite
Ca2+
-
strong inhibition
Co2+
-
strong inhibition
Cu2+
-
strong inhibition
Fe2+
-
strong inhibition
Fe3+
-
strong inhibition
Hg2+
-
strong inhibition
iodoacetic acid
-
-
iodonium diphenyl chloride
-
acts on NADPH- and FAD-binding domain of flavoprotein component
KCN
-
complete inhibition at 1 mM
Low ionic strength
-
NADPH-dependent activities
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o-phenanthroline
-
-
p-chloromercuribenzoate
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NADPH-dependent activities
p-Mercuriphenylsulfonate
PCMB
-
complete inhibition at 1 mM
sulfate
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-
Sulfide
additional information
-
inhibitory potency of metal ions, overview, no inhibition by Na+, K+, and Mg2+
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.06
cytochrome c
-
-
0.08 - 0.083
FAD
0.06 - 0.063
FMN
4.5 - 10.5
hydroxylamine
0.005 - 0.08
NADPH
0.8 - 1.5
nitrite
0.07
paraquat
-
-
0.021
reduced F420
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at a fixed sulfite concentration of 0.29 mM and within a reduced F420 concentration range of 0.004-0.06 mM
0.0043 - 2
sulfite
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
150
FAD
Escherichia coli
-
cosubstrate NADPH
98.3 - 217
hydroxylamine
Escherichia coli
-
cosubstrate NADPH
605
NADP+
Escherichia coli
-
-
1.25 - 468
NADPH
43.3 - 51.7
nitrite
Escherichia coli
-
cosubstrate NADPH
605
reduced methyl viologen
Escherichia coli
-
-
30 - 65
sulfite
additional information
additional information
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.08 - 0.12
arsenite
-
-
0.006 - 0.009
cyanide
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-
0.018
iodonium diphenyl chloride
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-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.0013
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in 50 mM potassium phosphate buffer (pH 7), 0.044 mM reduced F420, and 1.5 mM sodium sulfite, from cell extract
0.0182
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in 50 mM potassium phosphate buffer (pH 7), 0.044 mM reduced F420, and 1.5 mM sodium sulfite, after 14fold purification
2.73 - 2.87
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-
10.02
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purified enzyme
additional information
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-
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7 - 7.5
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NADPH-sulfite reductase
7.2
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electron donor NADPH
7.5 - 7.9
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NADPH-cytochrome c reductase
7.7 - 8.5
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methyl viologen-sulfite reductase
7.9
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sulfite
8.6
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nitrite
9.5
-
hydroxylamine
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.5 - 8
-
75% activity at pH 6.5 and pH 8.0
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
60000
-
determined by SDS-PAGE
119000
-
gel filtration
200000 - 208000
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gel filtration
300000
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gel filtration, sedimentation analysis at low ionic strength, dissociated
350000
488000
-
sedimentation under nondenaturing conditions
604000
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gel filtration
650000
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gel filtration, sedimentation analysis
670000
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sedimentation equilibrium centrifugation, sedimentation and diffusion coefficients
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
-
x * 60000, determined by SDS-PAGE
dodecamer
heterotetramer
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two alpha- and two beta-subunits, encoded by the MET10 and MET5 gene, respectively
hexamer
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alpha2,beta2,gamma2, 2 * 50000 + 2 * 42000 + 2 * 12500, SDS-PAGE
tetramer
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alpha2,beta2, 2 * 116000 + 2 * 167000, SDS-PAGE
additional information
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
FNR-like domain of flavoprotein
-
hemoprotein subunit
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hemoprotein subunit; x-ray structure of hemoprotein subunit
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recombinant monomeric fragment of flavoprotein, having 3 domains and binding FAD and NADPH
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.5 - 8
-
75% activity at pH 6.5 and pH 8.0 after 30 min at 25°C
656421
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50
-
inactivation above
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
low ionic strength such as 0.01 M phosphate buffer destroys NADPH-dependent activities
-
ORGANIC SOLVENT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Acetone
-
inactivation
Ethanol
-
inactivation
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-15°C, 0.05 M potassium phosphate buffer pH 7.7, 0.1 mM EDTA, 2 years stable
-
-15°C, 0.05 M potassium phosphate buffer pH 7.7, 10 mM EDTA, at least 6 months
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-15°C, reduced hemoprotein subunit or reduced flavoprotein subunit
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-20°C, 0.3 M phosphate buffer pH 7.3, 1 year stable stable
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4°C, 0.05 M potassium phosphate buffer pH 7.7, 0.1 mM EDTA, 1 month stable
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4°C, 0.3 M phosphate buffer pH 7.3, 1 week stable
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
579.5fold to homogeneity by ammoniums sulfate fractionation, DEAE ion exchange chromatography, and gel filtration
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phenyl-Sepharose chromatography, F420-Sepharose chromatography and QAE-Sephadex gel filtration
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preparation of FMN depleted enzyme
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preparation of mutant form lacking FAD and hence with no NADPH-sulfite reductase activity but with methyl viologen-sulfite reductase activity
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using a His-Trap column
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var. ellipsoideus (partial)
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
alpha and beta subunits
-
FMN-binding domain of flavoprotein component
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homology to cysI of Escherichia coli, i.e. alpha subunit
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NADPH- and FAD-binding domain of flavoprotein component
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the MET5 and Met10 coding sequences are cloned into the vector pGEM-T-easy and subsequently into the centromeric Saccharomyces cerevisiae expression plasmid pRS416
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the sulfite reductase hemo-subunit is cloned into the vector pLM1 for expression in Escherichia coli BL21DE3 cells
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C427A
-
mutation of the highly conserved cysteine 427, crucial residue for iron-sulfur cluster binding
C433A
-
mutation of the highly conserved cysteine 427, crucial residue for iron-sulfur cluster binding
C472A
-
mutation of the highly conserved cysteine 427, crucial residue for iron-sulfur cluster binding
C476A
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mutation of the highly conserved cysteine 427, crucial residue for iron-sulfur cluster binding
A979T
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Met5p mutant
E1356K
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Met5p mutant
E929K
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Met10p mutant
G1115D
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Met5p mutant
L606F
-
Met10p mutant
T990I
-
Met10p mutant
T997I
-
Met10p mutant
W59X
-
Met10p mutant
W841X
-
Met10p mutant
additional information
-
4 mutants blocked in sulfite reduction, 3 containing only FMN, 1 lacking both flavins
Renatured/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
mixing isolated hemoprotein and flavoprotein in appropriate proportions reconstituts activity
-
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
brewing
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mutations in the sulfite reductase genes, MET5 and MET10, are responsible for low H2S phenotypes, yeast strains with reduced H2S production offer promising solutions to H2S-related taints generated during wine production
medicine
-
potential use of vaccine against Actinobacillus pleuropneumoniae infections
nutrition
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quality determination of surimi, purified enzyme increases the reactive SH and gel strength of surimi prepared from frozen mackerel, processing of surimi-based products
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