Information on EC 1.2.7.11 - 2-oxoacid oxidoreductase (ferredoxin)

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
1.2.7.11
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RECOMMENDED NAME
GeneOntology No.
2-oxoacid oxidoreductase (ferredoxin)
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
a 2-oxocarboxylate + CoA + 2 oxidized ferredoxin = an acyl-CoA + CO2 + 2 reduced ferredoxin + 2 H+
show the reaction diagram
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Biosynthesis of antibiotics
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Butanoate metabolism
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Citrate cycle (TCA cycle)
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Glycolysis / Gluconeogenesis
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Metabolic pathways
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Microbial metabolism in diverse environments
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Pyruvate metabolism
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isoleucine metabolism
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threonine metabolism
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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).
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
metabolism
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-oxo-4-methylthiobutyrate + CoA + 2 methyl viologen
3-methylthiopropanoyl-CoA + CO2 + 2 reduced methyl viologen + 2 H+
show the reaction diagram
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enzyme Ape1473/1472, 48% of the activity compared to glyoxylate
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?
2-oxoadipate + CoA + 2 methyl viologen
glutaryl-CoA + CO2 + 2 reduced methyl viologen + 2 H+
show the reaction diagram
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enzyme Ape1473/1472, 49% of the activity compared to glyoxylate
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?
2-oxobutanoate + CoA + 2 oxidized cytochrome c
propanoyl-CoA + CO2 + 2 reduced cytochrome c + 2 H+
show the reaction diagram
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?
2-oxobutyrate + CoA + 2 oxidized methyl viologen
propanoyl-CoA + CO2 + 2 reduced methyl viologen
show the reaction diagram
2-oxobutyrate + CoA + oxidized ferredoxin
propanoyl-CoA + CO2 + reduced ferredoxin
show the reaction diagram
2-oxoglutarate + CoA + 2 oxidized cytochrome c
succinyl-CoA + CO2 + 2 reduced cytochrome c + 2 H+
show the reaction diagram
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best substrate tested
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?
2-oxoglutarate + CoA + 2 oxidized ferredoxin
succinyl-CoA + CO2 + 2 reduced ferredoxin + 2 H+
show the reaction diagram
2-oxoglutarate + CoA + 2 oxidized methyl viologen
succinyl-CoA + CO2 + 2 reduced methyl viologen + 2 H+
show the reaction diagram
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enzyme Ape1473/1472, 94% of the activity compared to glyoxylate
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?
4-hydroxyphenylpyruvate + CoA + 2 methyl viologen
(4-hydroxyphenyl)acetyl-CoA + CO2 + 2 reduced methyl viologen + 2 H+
show the reaction diagram
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enzyme Ape1473/1472, 16% of the activity compared to glyoxylate
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?
glyoxylate + CoA + 2 oxidized methyl viologen
formyl-CoA + CO2 + 2 reduced methyl viologen + 2 H+
show the reaction diagram
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enzyme Ape1473/1472
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?
hydroxypyruvate + CoA + 2 methyl viologen
?
show the reaction diagram
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enzyme Ape1473/1472, 55% of the activity compared to glyoxylate
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phenylpyruvate + CoA + 2 methyl viologen
phenylacetyl-CoA + CO2 + 2 reduced methyl viologen + 2 H+
show the reaction diagram
pyruvate + CoA + 2 oxidized cytochrome c
acetyl-CoA + CO2 + 2 reduced cytochrome c + 2 H+
show the reaction diagram
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?
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
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
pyruvate + CoA + 2 oxidized ferredoxin
acetyl-CoA + CO2 + 2 reduced ferredoxin + 2 H+
show the reaction diagram
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?
additional information
?
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Ape1473/1472 operates in the TCA cycle of Aeropyrum pernix
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Ferredoxin
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iron-sulfur centre
P72578 and P72579
the enzyme contains one [4Fe-4S]2+,1+ cluster
thiamine diphosphate
[4Fe-4S]-center
Q96XT2 and Q96XT4, Q96Y66 and Q96Y68
alpha/beta-subunit heterodimers contain 4Fe-4S cluster; alpha/beta-subunit heterodimers contain 4Fe-4S cluster
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4Fe-4S center
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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
iron-sulfur centre
P72578 and P72579
the enzyme contains one [4Fe-4S]2+,1+ cluster
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
4-fluoro-7-nitrobenzofurazan
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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
CoA
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substrate inhibition
KCl
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10–20% inhibition is observed with 50 mM KCl, while with higher concentrations (maximum, 0.6 M), 46–65% activation is reached, and further increased concentrations of KCl are inhibitory, enzyme Ape1473/1472
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.516
2-oxobutanoate
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pH 7.0, 55°C
0.07 - 0.516
2-oxobutyrate
0.163 - 15
2-oxoglutarate
0.05 - 1.17
CoA
0.07
oxidized ferredoxin
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cosubstrates: 1 mM 2-oxoxbutyrate, 0.05 mM CoA, pH and temperature not specified in the publication; cosubstrates: 1 mM pyruvate, 0.05 mM CoA, pH and temperature not specified in the publication
0.1 - 1.6
pyruvate
additional information
additional information
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kinetic parameters of mutant enzymes
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.5
2-oxobutanoate
Sulfolobus solfataricus
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pH 7.0, 55°C
1.55
2-oxobutyrate
Sulfolobus solfataricus
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pH 7.0, 55°C
1.7 - 19
2-oxoglutarate
2.2 - 51
pyruvate
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
3
2-oxobutanoate
Sulfolobus solfataricus
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pH 7.0, 55°C
390
3
2-oxobutyrate
Sulfolobus solfataricus
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pH 7.0, 55°C
371
0.52 - 46.2
2-oxoglutarate
34
7.2 - 182
pyruvate
31
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
52.6
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pH 7.0, 55°C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.8
P72578 and P72579
assay at
7.5
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assay at
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 9
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pH 6.0: about 50% of maximal activity, pH 9.0: about 50% of maximal activity
7.5 - 10
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pH 7.5: about 70% of maximal activity, pH 10.0: about 70% of maximal activity, enzyme Ape1473/1472
8 - 9
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pH 8.0: 65% of maximal activity, pH 9.0: optimum, inactive below pH 7.0
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50
P72578 and P72579
assay at
110
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enzyme Ape1473/1472
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25 - 52
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the enzyme is twice as active at 52°C as at 25°C
90 - 110
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70°C: 17% of maximal activity, 90°C: about 40% of maximal activity, 110°C: maximal activity, enzyme Ape1473/1472
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
34410
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1 * 67405 + 1 * 34410, calculated from sequence, enzyme Ape1473/1472
37000
P72578 and P72579
1 * 70000 (alpha) + 1 * 37000 (beta)
42000
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2 * 86000 (alpha) + 2 * 42000 (beta), SDS-PAGE
67000
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1 * 67000 + 1 * 34000, SDS-PAGE, enzyme Ape1473/1472
67405
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1 * 67405 + 1 * 34410, calculated from sequence, enzyme Ape1473/1472
69000
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1 * 69000, alpha-subuit, + 1 * 34000, beta-subunit, SDS-PAGE and MALDI-TOF; 1 * 69000 (alpha-subunit) + 1 * 34000 (beta-subunit), SDS-PAGE, MALDI-TOF mass spectrometry
70000
P72578 and P72579
1 * 70000 (alpha) + 1 * 37000 (beta)
71000
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1 * 71000 (subunit a) + 1 * 34000 (subunit b), SDS-PAGE
86000
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2 * 86000 (alpha) + 2 * 42000 (beta), SDS-PAGE
100000
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gel filtration, enzyme Ape1473/1472
103000
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native PAGE
105000
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gel filtration
165000
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sedimentation velocity experiments
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
heterodimer
tetramer
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2 * 86000 (alpha) + 2 * 42000 (beta), SDS-PAGE
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
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
Q96XT2 and Q96XT4, Q96Y66 and Q96Y68
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
70
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pH 7.2, 30 min, about 20% loss of activity, natural and recombinant wild-type enzyme
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
the enzymes requires high salt concentrations for stability. The half-life at 0.1 M KCl in 50 mM Tris/HC1 pH 8.0 is 3 h at 0°C
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OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
purified enzyme is stable against oxygen
P72578 and P72579
288435
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-80°C, purified enzyme can be stored after dialysis against 20 mM potassium phosphate buffer, pH 6.8
P72578 and P72579
20°C, 4°C or -80°C, activity in cytosolic fraction is not very stable
P72578 and P72579
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
;
Q96XT2 and Q96XT4, Q96Y66 and Q96Y68
enzyme Ape1473/1472
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wild-type and mutant enzymes
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
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P72578 and P72579
expression in Escherichia coli (DE3); expression in Escherichia coli (DE3)
Q96XT2 and Q96XT4, Q96Y66 and Q96Y68
expression in Escherichia coli, enzyme Ape1473/1472
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expression in Escherichia coli, wild-type recombinant enzyme is indistinguishable from the natural one in every criterion investigated
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expression of mutant enzymes K125A and K173A in Escherichia coli
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
I255L
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kcat/Km for pyruvate is 11% of wild-type value, kcat/KM for 2-oxoglutarate is 21% of wild-type value
I255M
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kcat/Km for pyruvate is 13% of wild-type value, kcat/KM for 2-oxoglutarate is 2% of wild-type value
I255S
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kcat/Km for pyruvate is 23% of wild-type value, kcat/KM for 2-oxoglutarate is 35% of wild-type value
I255V
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kcat/Km for pyruvate is 14% of wild-type value, kcat/KM for 2-oxoglutarate is 38% of wild-type value
P254G
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kcat/Km for pyruvate is 12% of wild-type value, kcat/KM for 2-oxoglutarate is 20% of wild-type value
P257A
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no enzyme activity at either 50 or 80°C
P257G
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no enzyme activity at either 50 or 80°C
P257V
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no enzyme activity at either 50 or 80°C
T256A
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kcat and Km for 2-oxoglutarate are 33% and 51%, respectively, as compared with that of the wild-type enzyme; kcat/Km for pyruvate is 21% of wild-type value, kcat/KM for 2-oxoglutarate is 15% of wild-type value
T256S
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kcat/Km for pyruvate is 15% of wild-type value, kcat/KM for 2-oxoglutarate is 92% of wild-type value
T256V
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kcat/Km for pyruvate is 17% of wild-type value, kcat/KM for 2-oxoglutarate is 68% of wild-type value
Y253A
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no enzyme activity at either 50 or 80°C
Y253F
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kcat/Km for pyruvate is 3.9% of wild-type value, kcat/KM for 2-oxoglutarate is 16% of wild-type value
Y253W
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no enzyme activity at either 50 or 80°C
T256A
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kcat and Km for 2-oxoglutarate are 33% and 51%, respectively, as compared with that of the wild-type enzyme; kcat/Km for pyruvate is 21% of wild-type value, kcat/KM for 2-oxoglutarate is 15% of wild-type value
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T256S
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kcat/Km for pyruvate is 15% of wild-type value, kcat/KM for 2-oxoglutarate is 92% of wild-type value
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T256V
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kcat/Km for pyruvate is 17% of wild-type value, kcat/KM for 2-oxoglutarate is 68% of wild-type value
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Y253F
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kcat/Km for pyruvate is 3.9% of wild-type value, kcat/KM for 2-oxoglutarate is 16% of wild-type value
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C12/15A
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loss of iron–sulfur cluster
C12A
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loss of iron–sulfur cluster. The mutant enzyme does not show formation of any radical intermediate or production of acetyl-CoA
C15A
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loss of iron–sulfur cluster. The mutant enzyme does not show formation of any radical intermediate or production of acetyl-CoA
C197A
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the enzyme retains an unidentified type of iron–sulfur cluster
C46A
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loss of iron–sulfur cluster. The mutant enzyme does not show formation of any radical intermediate or production of acetyl-CoA
D468A
Q96XT2 and Q96XT4, Q96Y66 and Q96Y68
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
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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
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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
Q96XT2 and Q96XT4, Q96Y66 and Q96Y68
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
Q96XT2 and Q96XT4, Q96Y66 and Q96Y68
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
Q96XT2 and Q96XT4, Q96Y66 and Q96Y68
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
D468A
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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
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K49I
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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
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S41A
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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
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T349L
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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
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