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Information on EC 7.2.1.3 - ascorbate ferrireductase (transmembrane) and Organism(s) Mus musculus and UniProt Accession Q9WUE3
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7.2.1.3 ascorbate ferrireductase (transmembrane)IUBMB Comments
A diheme cytochrome that transfers electrons across a single membrane, such as the outer membrane of the enterocyte, or the tonoplast membrane of the plant cell vacuole. Acts on hexacyanoferrate(III) and other ferric chelates.
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Word Map
- 7.2.1.3
- photosystem
- hepcidin
-
ferroportin
- hephaestin
-
iron-deficient
-
fe-deficient
- beta-hydroxylase
- hemochromatosis
-
high-potential
- monodehydroascorbate
-
oxygen-evolving
-
extravesicular
- pheophytin
-
iron-related
-
calcareous
-
ascorbate-reducible
-
flash-induced
- semidehydroascorbate
-
alpha-bands
-
3-3,4-dichlorophenyl-1,1-dimethylurea
- ferroxidase
-
fe-sufficient
The taxonomic range for the selected organisms is: Mus musculus
The enzyme appears in selected viruses and cellular organisms
The enzyme appears in selected viruses and cellular organisms
Reaction Schemes
Synonyms
cytochrome b561, cytochrome b-559, dcytb, ferric chelate reductase, duodenal cytochrome b, cybrd1, cyt b561, cyb561, cyt-b561, lcytb, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
ascorbate-cytochrome b5 reductase
-
-
-
-
CGCytb
DCytb
LCytb
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ascorbate[side 1] + Fe(III)[side 2] = monodehydroascorbate[side 1] + Fe(II)[side 2]
reaction mechanism
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
redox reaction
-
-
-
-
oxidation
-
-
-
-
reduction
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
Fe(III):ascorbate oxidorectuctase (electron-translocating)
A diheme cytochrome that transfers electrons across a single membrane, such as the outer membrane of the enterocyte, or the tonoplast membrane of the plant cell vacuole. Acts on hexacyanoferrate(III) and other ferric chelates.
CAS REGISTRY NUMBER
COMMENTARY
37237-57-3
-
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
ascorbate[side 1] + Fe3+-EDTA[side 2]
monodehydroascorbate[side 1] + ?
-
-
-
-
?
ascorbate[side 1] + Fe3+-EDTA[side 2]
monodehydroascorbate[side 1] + Fe2+-EDTA[side 2]
-
compared with FeCN, Fe3+-EDTA is a relatively poor substrate for the enzyme (20-35fold lower ferrireductase activities)
-
-
?
ascorbate[side 1] + ferricyanide[side 2]
monodehydroascorbate[side 1] + ferrocyanide[side2]
-
-
-
-
-
bathocuprionedisulfonate[side 1] + Cu(II)-nitrilotriacetic acid[side 2]
Cu(I)-bathocuproinedisulfonate
-
-
-
-
?
ferrozine[side 1] + Fe(III)-nitrilotriacetic acid[side 2]
Fe(II)-ferrozine
-
-
-
-
?
L-ascorbate + ferricytochrome b5
monodehydro-L-ascorbate + ferrocytochrome b5
-
-
-
-
?
ascorbate[side 1] + Fe(III)[side 2]
monodehydroascorbate[side 1] + Fe(II)[side 2]
-
-
-
-
?
ascorbate[side 1] + Fe(III)[side 2]
monodehydroascorbate[side 1] + Fe(II)[side 2]
-
-
-
?
additional information
?
-
-
Dcytb has the capacity to reduce both iron and copper complexes
-
-
?
additional information
?
-
the other heme-b center is responsible for the ascorbate oxidation by iozyme CGCytb. Structure-function relationship, overview
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
ascorbate[side 1] + Fe(III)[side 2]
monodehydroascorbate[side 1] + Fe(II)[side 2]
-
-
-
-
?
ascorbate[side 1] + Fe(III)[side 2]
monodehydroascorbate[side 1] + Fe(II)[side 2]
-
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
heme b
two heme-b centers and CYB561 protein, structure analysis and comparisons, overview. Midpoint redox potentials, spin, and spectra of heme b, comparisons
additional information
neither ferrocyanide nor durohydroquinone can reduce nCGCytb
-
ascorbate
cytosolic ascorbate is the cellular electron donor for the CYB561 proteins
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
dehydroascorbate
-
preloading cells with dehydroascorbate greatly increases both ferric and cupric reductase activities of Dcytb
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0152
Cu(II)-nitrilotriacetic acid[side 2]
-
in 25 mM MOPS, 25 mM MES, 5.4 mM KCl, 5 mM glucose, 140 mM NaCl, 1.8 mM CaCl2, 0.8 mM MgCl2, pH 7.0, at 22°C
0.0231
Cu(II)-nitrilotriacetic acid[side 2]
-
in 25 mM MOPS, 25 mM MES, 5.4 mM KCl, 5 mM glucose, 140 mM NaCl, 1.8 mM CaCl2, 0.8 mM MgCl2, pH 7.0, at 37°C
0.074
Fe(III)-nitrilotriacetic acid[side 2]
-
in 25 mM MOPS, 25 mM MES, 5.4 mM KCl, 5 mM glucose, 140 mM NaCl, 1.8 mM CaCl2, 0.8 mM MgCl2, pH 7.0, at 37°C
0.0921
Fe(III)-nitrilotriacetic acid[side 2]
-
in 25 mM MOPS, 25 mM MES, 5.4 mM KCl, 5 mM glucose, 140 mM NaCl, 1.8 mM CaCl2, 0.8 mM MgCl2, pH 7.0, at 22°C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
the rate of reduction of CYB561 by ascorbate is only slightly pH-dependent, steady-state reduction kinetics
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
CGCytb, the chromaffin granule CYB561 of the mammalian adrenal glands (CGCytb) makes up about 10-15% of the adrenal gland chromaffin granule membrane proteins
a transmembrane enzyme, localization of both the N- and C-termini of CGCytb in the cytoplasm. Native CGCytb is a transmembrane electron transferring protein that has 6 transmembrane domains with two pairs of His residues, arranged on four consecutive transmembrane domains (the CYB561-core), for coordinating two b-type hemes, one on each side of the membrane, transmembrane orientation, modeling
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highly expressed in the brush-border membrane of duodenal enterocytes
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
metabolism
enzyme is reduced by dihydrolipoic acid almost as efficiently as by ascorbate
malfunction
mutation of His residues coordinating the intra-vesicular-side heme-b results in an almost complete loss of protein, while mutation of His residues coordinating the cytosolic-side heme-b hardly affects the expression of CYB561 proteins but results in a changed reducibility and heme content of these proteins. Replacing any of the 4 highly conserved His residues, coordinating the two b-type hemes, by Ala in mouse rLCytb completely abolishes the transmembrane ferric reductase activity of rLCytb
physiological function
additional information
the binding sites for the ascorbate on the cytoplasmic and the monodehydroascorbate on the non-cytoplasmic side do not correspond to the putative binding sites that had been inferred from the sequence (homology) analysis as well as from site directed mutagenesis of a number of CYB561 proteins. Models for the sidedness of CYB561 enzymes and the reduction by ascorbate, overview. Importance of an Arg residue in the reduction of rCGCytb
physiological function
-
Dcytb plays a physiological role in both iron and copper uptake, through divalent metal transporter1 and copper transporter 1, respectively
physiological function
b-type cytochromes are heme-containing, electron-transporting proteins in which the redox active center(s) is (are) iron-protoporphyrin(s) IX non-covalently bound to the protein matrix. Some of the b-type cytochromes are localized in membranous structures and have two heme-b prosthetic groups, the major function of these proteins is transmembrane electron transport. Isozyme DCytb is capable of reducing ferric chelates and plays an important role in the iron acquisition of cells
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). C56D2_MOUSE
222
6
24187
Swiss-Prot
Secretory Pathway (Reliability: 5)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
Cytb561 enzyme structure analysis, structure-function relationship, detailed overview
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
D38A
E117A
-
the mutant shows decreased Fe(CN)3 reductase activity compared to the wild type enzyme
E177A
-
the mutation of lysosomal cytochrome b561 reduces the enzyme activity compared to the wild type
E196A
F44A
H105A
H108A
site-directed mutagenesis, the mutation results in a practically unchanged level of protein expression and a considerably lower ascorbate reducibility
H112A
H117A
H120A
site-directed mutagenesis, the mutation results in nearly undetectable levels of rCGCytb
H156A
H159A
site-directed mutagenesis, the mutation results in a practically unchanged level of protein expression and a considerably lower ascorbate reducibility
H47A
H52A
site-directed mutagenesis, the mutation results in nearly undetectable levels of rCGCytb
H83A
H86A
site-directed mutagenesis, the mutation results in a practically unchanged level of protein expression and a considerably lower ascorbate reducibility
H86A/H159A
site-directed mutagenesis, the mutation results in a practically unchanged level of protein expression and a considerably lower ascorbate reducibility
M51A
N106A
P48A
Q131A
R149A
R67A
R72A
site-directed mutagenesis, the mutant shows reduced activity compared to wild-type
R72E
R72K
site-directed mutagenesis, the mutant shows reduced activity compared to wild-type
R72T
site-directed mutagenesis, the mutant shows reduced activity compared to wild-type
R72Y
site-directed mutagenesis, the mutant shows reduced activity compared to wild-type
S115A
S118A
-
the mutation of lysosomal cytochrome b561 increases the enzyme activity compared to the wild type
W119A
Y190A
Y66A
additional information
replacing any of the 4 highly conserved His residues, coordinating the two b-type hemes, by Ala in mouse rLCytb completely abolishes the transmembrane ferric reductase activity of rLCytb. Midpoint ascorbate concentration for the reduction of low-potential heme-b centers is hardly influenced by the R74X replacements but that for the high-potential heme-b centers show a significant trend
D38A
-
the mutant shows increased Fe(CN)3 reductase activity compared to the wild type enzyme
D38A
-
the mutation of lysosomal cytochrome b561 increases the enzyme activity compared to the wild type
E196A
-
the mutant shows decreased Fe(CN)3 reductase activity compared to the wild type enzyme
E196A
-
the mutation of lysosomal cytochrome b561 strongly reduces the enzyme activity compared to the wild type
F44A
-
the mutation of lysosomal cytochrome b561 reduces the enzyme activity by about 45% compared to the wild type
H105A
-
the mutant shows increased Fe(CN)3 reductase activity compared to the wild type enzyme
H105A
-
the mutation of lysosomal cytochrome b561 increases the enzyme activity compared to the wild type
H112A
-
the mutant shows decreased Fe(CN)3 reductase activity compared to the wild type enzyme
H112A
-
the mutation of lysosomal cytochrome b561 reduces the enzyme activity compared to the wild type
H117A
-
the mutation completely abrogates the Fe(CN)3 reductase activity of the enzyme
H117A
-
the mutation of lysosomal cytochrome b561 completely abrogates the FeCN reductase activity of the enzyme
H156A
-
the mutation completely abrogates the Fe(CN)3 reductase activity of the enzyme
H156A
-
the mutation of lysosomal cytochrome b561 completely abrogates the FeCN reductase activity of the enzyme
H47A
-
the mutation completely abrogates the Fe(CN)3 reductase activity of the enzyme
H47A
-
the mutation of lysosomal cytochrome b561 completely abrogates the FeCN reductase activity of the enzyme
H83A
-
the mutation completely abrogates the Fe(CN)3 reductase activity of the enzyme
H83A
site-directed mutagenesis, no alteration is found from the ascorbate reducibility compared to mouse wild-type rCGCytb
H83A
-
the mutation of lysosomal cytochrome b561 completely abrogates the FeCN reductase activity of the enzyme
M51A
-
the mutant shows increased Fe(CN)3 reductase activity compared to the wild type enzyme
M51A
-
the mutation of lysosomal cytochrome b561 increases the enzyme activity compared to the wild type
N106A
-
the mutant shows increased Fe(CN)3 reductase activity compared to the wild type enzyme
N106A
-
the mutation of lysosomal cytochrome b561 increases the enzyme activity compared to the wild type
P48A
-
the mutant shows increased reductase activity compared to the wild type enzyme
P48A
-
the mutation of lysosomal cytochrome b561 increases the enzyme activity compared to the wild type
Q131A
-
the activity of the mutant is reduced significantly to 45% of that of the wild type
Q131A
-
the mutation of lysosomal cytochrome b561 reduces the enzyme activity to 45% of that of the wild type
R149A
-
the mutation results in a 75% loss in activity compared to the wild type enzyme
R149A
-
the mutation of lysosomal cytochrome b561 results in a 75% loss in activity compared to the wild type enzyme
R67A
-
the mutation results in an almost complete loss of the Fe(CN)3 reductase activity
R67A
-
the mutation of lysosomal cytochrome b561 results in an almost complete loss of the FeCN reductase activity of the enzyme
R72E
site-directed mutagenesis, the mutant shows reduced activity compared to wild-type
R72E
site-directed mutagenesis, the mutation of TCytb does not affect the final reduction level of rTCytb by ascorbate but results in a complete loss of the pH-dependent initial time-lag upon electron acceptance from ascorbate
S115A
-
the ferrireductase activity of lysosomal cytochrome b561 in mutant S115A is reduced to 50% compared to the wild type
W119A
-
the mutant shows decreased Fe(CN)3 reductase activity compared to the wild type enzyme
W119A
-
the ferrireductase activity of lysosomal cytochrome b561 in mutant W119A is reduced to 17% compared to the wild type
Y190A
-
the mutant shows increased Fe(CN)3 reductase activity compared to the wild type enzyme
Y190A
-
the mutation of lysosomal cytochrome b561 increases the enzyme activity compared to the wild type
Y66A
-
the mutation results in an almost complete loss of the Fe(CN)3 reductase activity
Y66A
-
the mutation of lysosomal cytochrome b561 results in an almost complete loss of the FeCN reductase activity of the enzyme
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant C-terminaly His6-tagged enzyme from yeast YPH499 cells by nickel affinity chromatography, recombinant C-terminally His6-tagged isozyme DCytb from Escherichia coli
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expressed in a Saccharomyces cerevisiae DELTAfre1DELTAfre2 mutant, which lacks almost all of its plasma membrane ferrireductase activity
-
expressed in a Saccharomyces cerevisiae strain S288C DELTAfre1DELTAfre2 mutant, which lacks almost all of its plasma membrane ferrireductase activity
-
recombinant expression of C-terminaly His6-tagged enzyme in yeast YPH499 cells, recombinant expression of C-terminally His6-tagged isozyme DCytb in Escherichia coli
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Knoepfel, M.; Solioz, M.
Characterization of a cytochrome b558 ferric/cupric reductase from rabbit duodenal brush border membranes
Biochem. Biophys. Res. Commun.
291
220-225
2002
Oryctolagus cuniculus, Mus musculus
Su, D.; Asard, H.
Three mammalian cytochromes b561 are ascorbate-dependent ferrireductases
FEBS J.
273
3722-3734
2006
Homo sapiens, Mus musculus, Rattus norvegicus
Wyman, S.; Simpson, R.J.; McKie, A.T.; Sharp, P.A.
Dcytb (Cybrd1) functions as both a ferric and a cupric reductase in vitro
FEBS Lett.
582
1901-1906
2008
Mus musculus
McKie, A.T.; Barrow, D.; Latunde-Dada, G.O.; Rolfs, A.; Sager, G.; Mudaly, E.; Mudaly, M.; Richardson, C.; Barlow, D.; Bomford, A.; Peters, T.J.; Raja, K.B.; Shirali, S.; Hediger, M.A.; Farzaneh, F.; Simpson, R.J.
An iron-regulated ferric reductase associated with the absorption of dietary iron
Science
291
1755-1759
2001
Mus musculus
Berczi, A.; Zimanyi, L.; Asard, H.
Dihydrolipoic acid reduces cytochrome b561 proteins
Eur. Biophys. J.
42
159-168
2013
Arabidopsis thaliana (Q8L856), Mus musculus (Q9WUE3)
Berczi, A.; Zimanyi, L.
The trans-membrane cytochrome b561 proteins structural information and biological function
Curr. Protein Pept. Sci.
15
745-760
2014
Bos taurus (P10897), Homo sapiens (P49447), Schistosoma japonicum (Q5D8X4), Zea mays (Q6I681), Mus musculus (Q6P1H1), Arabidopsis thaliana (Q9SWS1)
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