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Information on EC 1.2.7.11 - 2-oxoacid oxidoreductase (ferredoxin) and Organism(s) Sulfurisphaera tokodaii and UniProt Accession Q96Y68

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IUBMB Comments
Contains thiamine diphosphate and [4Fe-4S] clusters . This enzyme is a member of the 2-oxoacid oxidoreductases, a family of enzymes that oxidatively decarboxylate different 2-oxoacids to form their CoA derivatives, and are differentiated based on their substrate specificity. For example, see EC 1.2.7.3, 2-oxoglutarate synthase and EC 1.2.7.7, 3-methyl-2-oxobutanoate dehydrogenase (ferredoxin).
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Sulfurisphaera tokodaii
UNIPROT: Q96Y68
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The taxonomic range for the selected organisms is: Sulfurisphaera tokodaii
The expected taxonomic range for this enzyme is: Archaea, Bacteria
Synonyms
2-oxoacid:ferredoxin oxidoreductase, 2-oxoacid:ferredoxin oxidoreductases, stofor, ape1473/1472, stofor2, stofor1, 2-oxoacid ferredoxin oxidoreductase, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
2-oxoacid:ferredoxin oxidoreductases
-
2-oxoacid:ferredoxin oxidoreductase
2-oxoacid:ferredoxin oxidoreductases
-
StOFOR1
StOFOR2
SYSTEMATIC NAME
IUBMB Comments
2-oxocarboxylate:ferredoxin 2-oxidoreductase (decarboxylating, CoA-acylating)
Contains thiamine diphosphate and [4Fe-4S] clusters [2]. This enzyme is a member of the 2-oxoacid oxidoreductases, a family of enzymes that oxidatively decarboxylate different 2-oxoacids to form their CoA derivatives, and are differentiated based on their substrate specificity. For example, see EC 1.2.7.3, 2-oxoglutarate synthase and EC 1.2.7.7, 3-methyl-2-oxobutanoate dehydrogenase (ferredoxin).
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
2-oxobutyrate + CoA + 2 oxidized methyl viologen
propanoyl-CoA + CO2 + 2 reduced methyl viologen
show the reaction diagram
-
-
-
?
2-oxoglutarate + CoA + oxidized ferredoxin
succinyl-CoA + CO2 + reduced ferredoxin + H+
show the reaction diagram
-
-
-
?
2-oxoglutarate + CoA + oxidized methyl viologen
succinyl-CoA + CO2 + reduced methyl viologen + H+
show the reaction diagram
-
-
-
?
pyruvate + CoA + oxidized methyl viologen
?
show the reaction diagram
-
-
-
?
2-oxobutyrate + CoA + 2 oxidized methyl viologen
propanoyl-CoA + CO2 + 2 reduced methyl viologen
show the reaction diagram
-
-
-
?
2-oxoglutarate + CoA + 2 oxidized ferredoxin
succinyl-CoA + CO2 + 2 reduced ferredoxin + 2 H+
show the reaction diagram
2-oxoglutarate + CoA + oxidized ferredoxin
succinyl-CoA + CO2 + reduced ferredoxin + H+
show the reaction diagram
-
-
-
?
2-oxoglutarate + CoA + oxidized methyl viologen
succinyl-CoA + CO2 + reduced methyl viologen + H+
show the reaction diagram
-
-
-
?
pyruvate + CoA + 2 oxidized ferredoxin
acetyl-CoA + CO2 + 2 reduced ferredoxin + 2 H+
show the reaction diagram
pyruvate + CoA + 2 oxidized methyl viologen
acetyl-CoA + CO2 + 2 reduced methyl viologen + 2 H+
show the reaction diagram
-
-
-
?
pyruvate + CoA + oxidized methyl viologen
?
show the reaction diagram
-
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
2-oxoglutarate + CoA + oxidized ferredoxin
succinyl-CoA + CO2 + reduced ferredoxin + H+
show the reaction diagram
-
-
-
?
2-oxoglutarate + CoA + oxidized ferredoxin
succinyl-CoA + CO2 + reduced ferredoxin + H+
show the reaction diagram
-
-
-
?
pyruvate + CoA + 2 oxidized ferredoxin
acetyl-CoA + CO2 + 2 reduced ferredoxin + 2 H+
show the reaction diagram
-
-
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Ferredoxin
alpha/beta-subunit heterodimers lack an intramolecular ferredoxin domain. Because 2-oxoacid oxidoreductase enzymes from Sulfolobus tokodaii lack the intramolecular Fd-like domain V, there is a large pocket surrounded by domains III and VI in each protomer, which appears to be able to bind an external ferredoxin molecule
-
thiamine diphosphate
[4Fe-4S]-center
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4Fe-4S center
-
the enzyme has one [4Fe–4S]2+ cluster, ligated by 4 Cys residues, C12, C15, C46, and C197. All four Cys are required to fold a [4Fe–4S] cluster for oxidative decarboxylation of pyruvate including the formation of a stable hydroxyethyl-thiamine diphosphate radical
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
4-fluoro-7-nitrobenzofurazan
-
0.001 mM inhibitor with 0.01 mM enzyme decreases the activity by about 20% and 0.003 mM inhibitor with 0.01 mM enzyme decreases the activity by around 50%. Inactivation is prevented by CoA
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2.1 - 3.2
2-oxoglutarate
0.32 - 0.91
pyruvate
2.1 - 15
2-oxoglutarate
0.05 - 1.17
CoA
0.32 - 1.6
pyruvate
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
beta-subunit
UniProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
metabolism
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
heterodimer
1 * 70000 + 1 * 34000, SDS-PAGE
heterodimer
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
sitting drop vapor diffusion method, using 15% (w/v) PEG3350, 0.1 M Tris-HCl (pH 8.5) and 0.1 M NaBr
crystals are grown at 20°C using sitting drop vapor diffusion. The structure of the recombinant enzyme StOFOR2 by the single-wavelength anomalous dispersion method is solved using a selenomethionine(SeMet)-labeled protein crystal, and the structures of the ligand-free (2.1 Å resolution) and pyruvate-complexed (2.2 Å) forms are determined. In the structure of the recombinant enzyme StOFOR2 in unreacted pyruvate complex form, the carboxylate group of pyruvate is recognized by Arg344 and Thr257 from the alpha-subunit. The binding pockets of the 2-oxoacid oxidoreductase enzymes from Sulfolobus tokodaii surrounding the methyl or propyl group of the ligands are wider than that of 2-oxoacid oxidoreductase enzymes from Desulfovbrio africanus. A possible complex structural model is constructed by placing a Zn2+-containing dicluster ferredoxin of Sulfolobus tokodaii into the large pocket of the recombinant StOFOR2 enzyme, providing insight into the electron transfer between the two redox proteins
crystals of the StOFOR1 enzyme are prepared by co-crystallization with 50 mM 2-oxobutyrate and 1 mM CoA. Crystals are grown at 25°C using sitting drop vapor diffusion. In the structure of StOFOR1 co-crystallized with 2-oxobutyrate, electron density corresponding to a 1-hydroxypropyl group (post-decarboxylation state) is observed at the thiazole ring of thiamine diphosphate. The binding pockets of the 2-oxoacid oxidoreductase enzymes from Sulfolobus tokodaii surrounding the methyl or propyl group of the ligands are wider than that of 2-oxoacid oxidoreductase enzymes from Desulfovbrio africanus
sitting drop vapor diffusion method, using 0.7 M ammonium tartrate dibasic and 0.1 M Tris-HCl (pH 8.5)
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D468A
inactive with 2-oxoglutarate and pyruvate as substrate
K49I
inactive with 2-oxoglutarate as substrate
S41A
the mutant shows reduced activity compared to the wild type enzyme
T349L
the mutant shows reduced activity compared to the wild type enzyme
C12/15A
-
loss of iron–sulfur cluster
C12A
-
loss of iron–sulfur cluster. The mutant enzyme does not show formation of any radical intermediate or production of acetyl-CoA
C15A
-
loss of iron–sulfur cluster. The mutant enzyme does not show formation of any radical intermediate or production of acetyl-CoA
C197A
-
the enzyme retains an unidentified type of iron–sulfur cluster
C46A
-
loss of iron–sulfur cluster. The mutant enzyme does not show formation of any radical intermediate or production of acetyl-CoA
D468A
mutant enzyme StOFOR1 with mutation D468A in alpha-subunit. Vmax with pyruvate as substrate is 1.3% compared to wild-type enzyme. No activity is detected with 2-oxoglutarate
K125A
-
the mutant enzyme shows a large increase in the Km-value for CoA and shows poor inactivation by 4-fluoro-7-nitrobenzofurazan, compared with K173A and wild type enzyme
K173A
-
the mutant enzyme shows a large increase in the Km-value for CoA and shows poor inactivation by 4-fluoro-7-nitrobenzofurazan, compared with K173A and wild type enzyme
K49I
mutant enzyme StOFOR1 with mutation K49I in alpha-subunit. Vmax with pyruvate as substrate is 28% compared to wild-type enzyme, Km with pyruvate as substrate is 2.8fold higher as compared to wild-type enzyme. No activity is detected with 2-oxoglutarate
S41A
mutant enzyme StOFOR1 with mutation S41A in alpha-subunit. Vmax with pyruvate as substrate is 29% compared to wild-type enzyme, Vmax with 2-oxoglutarate as substrate is 40% compared to wild-type enzyme, Km with pyruvate as substrate is 1.5fold higher as compared to wild-type enzyme, Km with 2-oxoglutarate as substrate is 1.5fold higher as compared to wild-type enzyme
T349L
mutant enzyme StOFOR1 with mutation T349L in alpha-subunit. Vmax with pyruvate as substrate is 43% compared to wild-type enzyme, Vmax with 2-oxoglutarate as substrate is 74% compared to wild-type enzyme, Km with pyruvate as substrate is 1.6fold higher as compared to wild-type enzyme, Km with 2-oxoglutarate as substrate is 1.1fold higher as compared to wild-type enzyme
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
DEAE Sepharose column chromatography and Superdex 200 gel filtration
DEAE Sepharose column chromatography and Superdex 200 gel filtration
wild-type and mutant enzymes
-
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli C43 (DE3) cells
expressed in Escherichia coli C43 (DE3) cells
expression in Escherichia coli (DE3)
expression of mutant enzymes K125A and K173A in Escherichia coli
-
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Luo, J.; Fukuda, E.; Takase, H.; Fushinobu, S.; Shoun, H.; Wakagi, T.
Identification of the lysine residue responsible for coenzyme A binding in the heterodimeric 2-oxoacid:ferredoxin oxidoreductase from Sulfolobus tokodaii, a thermoacidophilic archaeon, using 4-fluoro-7-nitrobenzofurazan as an affinity label
Biochim. Biophys. Acta
1794
335-340
2009
Sulfurisphaera tokodaii
Manually annotated by BRENDA team
Yan, Z.; Fushinobu, S.; Wakagi, T.
Four Cys residues in heterodimeric 2-oxoacid:ferredoxin oxidoreductase are required for CoA-dependent oxidative decarboxylation but not for a non-oxidative decarboxylation
Biochim. Biophys. Acta
1844
736-743
2014
Sulfurisphaera tokodaii
Manually annotated by BRENDA team
Yan, Z.; Maruyama, A.; Arakawa, T.; Fushinobu, S.; Wakagi, T.
Crystal structures of archaeal 2-oxoacid:ferredoxin oxidoreductases from Sulfolobus tokodaii
Sci. Rep.
6
33061
2016
Sulfurisphaera tokodaii (Q96XT2 and Q96XT4), Sulfurisphaera tokodaii (Q96XT4), Sulfurisphaera tokodaii (Q96Y66 and Q96Y68), Sulfurisphaera tokodaii (Q96Y68), Sulfurisphaera tokodaii, Sulfurisphaera tokodaii 7 (Q96XT4), Sulfurisphaera tokodaii 7 (Q96Y68), Sulfurisphaera tokodaii DSM 16993 (Q96XT2 and Q96XT4), Sulfurisphaera tokodaii DSM 16993 (Q96Y66 and Q96Y68)
Manually annotated by BRENDA team