Information on EC 4.1.1.72 - branched-chain-2-oxoacid decarboxylase

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

EC NUMBER
COMMENTARY
4.1.1.72
-
RECOMMENDED NAME
GeneOntology No.
branched-chain-2-oxoacid decarboxylase
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
(3S)-3-methyl-2-oxopentanoate = 2-methylbutanal + CO2
show the reaction diagram
acts on a number of 2-oxo acids, with a high affinity toward branched-chain substrates. The aldehyde formed may be enzyme-bound, and may be an intermediate in the bacterial system for the biosynthesis of branched-chain fatty acids
-
-
-
(3S)-3-methyl-2-oxopentanoate = 2-methylbutanal + CO2
show the reaction diagram
binding of cofactor thiamine diphosphate to E1b component of enzyme induces a disorder-to-order transition of the conserved phosphorylation loop carrying phosphorylation sites S292 and S302
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
decarboxylation
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
butanol and isobutanol biosynthesis (engineered)
-
-
L-isoleucine degradation II
-
-
L-valine degradation II
-
-
methionine metabolism
-
-
pyruvate fermentation to isobutanol (engineered)
-
-
SYSTEMATIC NAME
IUBMB Comments
(3S)-3-methyl-2-oxopentanoate carboxy-lyase (2-methylbutanal-forming)
Acts on a number of 2-oxo acids, with a high affinity towards branched-chain substrates. The aldehyde formed may be enzyme-bound, and may be an intermediate in the bacterial system for the biosynthesis of branched-chain fatty acids.
SYNONYMS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
BCKA
-
-
-
-
branched-chain 2-oxoacid decarboxylase
-
-
-
-
branched-chain alpha-keto acid decarboxylase
-
-
-
-
branched-chain keto acid decarboxylase
-
-
-
-
branched-chain oxo acid decarboxylase
-
-
-
-
decarboxylase, branched-chain oxo acid
-
-
-
-
CAS REGISTRY NUMBER
COMMENTARY
63653-19-0
-
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
expressed in Lycopersicon esculentum
-
-
Manually annotated by BRENDA team
simultaneous expression of human alpha-subunit and bovine beta-subunit of enzyme in Escherichia coli
-
-
Manually annotated by BRENDA team
simultaneous expression of human alpha-subunit and bovine beta-subunit of enzyme in Escherichia coli
-
-
Manually annotated by BRENDA team
subspecies Lactococcus lactis cremoris, strain B1157
-
-
Manually annotated by BRENDA team
enzyme may be a component of an enzyme complex named alpha-ketoisocaproate:alpha-keto-beta-methylvalerate dehydrogenase
-
-
Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                      
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
(indol-3-yl)pyruvate
(indol-3-yl)acetaldehyde + CO2
show the reaction diagram
P9WG37
-
-
-
?
2-oxo-3-methylpentanoate
2-methylbutanal + CO2
show the reaction diagram
P9WG37
-
-
-
?
2-oxo-3-methylvalerate
2-methylbutanal + CO2
show the reaction diagram
-
the enzymatic activity toward 2-oxo-3-methylvalerate is 1.5times higher than the activity toward 2-oxo-isovalerate
-
-
?
2-oxo-3-phenylpropanoic acid
phenylacetaldehyde + CO2
show the reaction diagram
-
at 30 mM 8.6% activity relative to 3-methyl-2-oxobutanoic acid
-
-
?
2-oxobutanoic acid
propanal + CO2
show the reaction diagram
Q6QBS4
10% of the rate with 3-methyl-2-oxobutanoic acid
-
-
?
2-oxobutanoic acid
propanal + CO2
show the reaction diagram
-
at 30 mM 9.3% activity relative to 3-methyl-2-oxobutanoic acid
-
-
?
2-oxobutyrate
propionaldehyde + CO2
show the reaction diagram
P9WG37
-
-
-
?
2-oxoglutarate
CO2 + succinate semialdehyde
show the reaction diagram
-
5% of the activity with L-3-methyl-2-oxopentanoate
-
-
-
2-oxohexanoate
n-pentanal + CO2
show the reaction diagram
P9WG37
-
-
-
?
2-oxohexanoic acid
pentanal + CO2
show the reaction diagram
Q6QBS4
30% of the rate with 3-methyl-2-oxobutanoic acid
-
-
?
2-oxohexanoic acid
pentanal + CO2
show the reaction diagram
-
at 30 mM 13% activity relative to 3-methyl-2-oxobutanoic acid
-
-
?
2-oxoisohexanoate
CO2 + isopentanal
show the reaction diagram
-
38% of the activity with L-3-methyl-2-oxopentanoate
-
-
-
2-oxoisohexanoate
CO2 + isopentanal
show the reaction diagram
-
38% of the activity with L-3-methyl-2-oxopentanoate
-
-
-
2-oxoisohexanoate
? + CO2
show the reaction diagram
P9WG37
-
-
-
?
2-oxoisopentanoate
CO2 + isobutanal
show the reaction diagram
-
63% of the activity with L-3-methyl-
-
-
?
2-oxoisopentanoate
2-methylpropanal + CO2
show the reaction diagram
P9WG37
-
-
-
?
2-oxoisovalerate
2-methylpropanal + CO2
show the reaction diagram
-
-
-
-
?
2-oxopentanoate
n-butanal + CO2
show the reaction diagram
P9WG37
-
-
-
?
2-oxopentanoic acid
butanal + CO2
show the reaction diagram
Q6QBS4
25% of the rate with 3-methyl-2-oxobutanoic acid
-
-
?
2-oxopentanoic acid
butanal + CO2
show the reaction diagram
-
at 30 mM 15% activity relative to 3-methyl-2-oxobutanoic acid
-
-
?
2-oxopropanoic acid
acetaldehyde + CO2
show the reaction diagram
-
at 30 mM 1.2% activity relative to 3-methyl-2-oxobutanoic acid
-
-
?
3-(4-hydroxyphenyl)-2-oxopropanoic acid
(4-hydroxyphenyl)-acetaldehyde + CO2
show the reaction diagram
-
at 30 mM 6% activity relative to 3-methyl-2-oxobutanoic acid
-
-
?
3-methyl-2-oxobutanoic acid
2-methylpropanal + CO2
show the reaction diagram
-
-
-
-
?
3-methyl-2-oxobutanoic acid
2-methylpropanal + CO2
show the reaction diagram
Q6QBS4
-
-
-
?
3-methyl-2-oxobutanoic acid
2-methylpropanal + CO2
show the reaction diagram
-
100% activity
-
-
?
3-methyl-2-oxopentanoate
CO2 + 2-methylbutanal
show the reaction diagram
-
-
-
-
?
3-methyl-2-oxopentanoate
CO2 + 2-methylbutanal
show the reaction diagram
-
-
-
-
?
3-methyl-2-oxopentanoic acid
2-methylbutanal + CO2
show the reaction diagram
Q6QBS4
30% of the rate with 3-methyl-2-oxobutanoic acid
-
-
?
3-methyl-2-oxopentanoic acid
2-methylbutanal + CO2
show the reaction diagram
-
at 30 mM 38.1% activity relative to 3-methyl-2-oxobutanoic acid
-
-
?
4-(4-hydroxyphenyl)-2-oxobutanoic acid
3-(4-hydroxyphenyl)-propanal + CO2
show the reaction diagram
-
at 30 mM 1.6% activity relative to 3-methyl-2-oxobutanoic acid
-
-
?
4-(methylsulfanyl)-2-oxobutanoic acid
3-(methylsulfanyl)-propanal + CO2
show the reaction diagram
-
at 30 mM 18% activity relative to 3-methyl-2-oxobutanoic acid
-
-
?
4-hydroxyphenylpyruvate
4-hydroxyphenylacetaldehyde + CO2
show the reaction diagram
P9WG37
-
-
-
?
4-methyl-2-oxohexanoic acid
3-methylpentanal + CO2
show the reaction diagram
-
at 30 mM 19% activity relative to 3-methyl-2-oxobutanoic acid
-
-
?
4-methyl-2-oxopentanoic acid
3-methylbutanal + CO2
show the reaction diagram
Q6QBS4
35% of the rate with 3-methyl-2-oxobutanoic acid
-
-
?
4-methyl-2-oxopentanoic acid
3-methylbutanal + CO2
show the reaction diagram
-
at 30 mM 26.3% activity relative to 3-methyl-2-oxobutanoic acid
-
-
?
4-methylthio-2-oxobutanoic acid
3-methylthiopropanal + CO2
show the reaction diagram
Q6QBS4
10% of the rate with 3-methyl-2-oxobutanoic acid
-
-
?
5-(4-hydroxyphenyl)-2-oxopentanoic acid
4-(4-hydroxyphenyl)-butanal + CO2
show the reaction diagram
-
at 30 mM 1.1% activity relative to 3-methyl-2-oxobutanoic acid
-
-
?
alpha-isocaproic acid
3-methylbutanal + CO2
show the reaction diagram
Q6QBS4
-
-
-
?
benzaldehyde + CO2
(R)-benzoin
show the reaction diagram
-
self-carboligation
-
-
?
benzoylformate
? + CO2
show the reaction diagram
P9WG37
-
-
-
?
indole-3-pyruvate
?
show the reaction diagram
-
at 1 mM 0.85% activity relative to 3-methyl-2-oxobutanoic acid
-
-
?
isovaleraldehyde
?
show the reaction diagram
-
self-carboligation
-
-
?
L-3-methyl-2-oxopentanoate
CO2 + 2-methylbutanal
show the reaction diagram
-
-
-
-
?
oxo(phenyl)acetic acid
benzaldehyde + CO2
show the reaction diagram
-
at 30 mM 8.4% activity relative to 3-methyl-2-oxobutanoic acid
-
-
?
Phenylpyruvate
Phenylacetaldehyde + CO2
show the reaction diagram
-
-
-
-
?
Phenylpyruvate
Phenylacetaldehyde + CO2
show the reaction diagram
P9WG37
-
-
-
?
Phenylpyruvate
Phenylacetaldehyde + CO2
show the reaction diagram
P9WG37
-
-
-
?
Phenylpyruvate
Phenylacetaldehyde + CO2
show the reaction diagram
-
-
-
-
?
Pyruvate
Acetaldehyde + CO2
show the reaction diagram
P9WG37
-
-
-
?
pyruvate
CO2 + ethanal
show the reaction diagram
-
25% of the activity with L-3-methyl-2-oxopentanoate
-
-
-
L-3-methyl-2-oxopentanoate
CO2 + 2-methylbutanal
show the reaction diagram
-
stereospecificity towards the L-isomer
-
-
?
additional information
?
-
-
essential for the synthesis of branched-chain fatty acids
-
-
-
additional information
?
-
-
plants with enzymic activity show enhanced cold tolerance, role as a protective mechanism for growth of plants under sub optimal temperatures
-
-
-
additional information
?
-
-
3-fluoro-2-oxopropanoic acid, isobutyraldehyde, 3,5-dichlorobenzaldehyde, 3,5-dimethoxybenzaldehyde and 2-oxooctanoic acid are no substrates
-
-
-
additional information
?
-
-
the enzyme is also able to catalyze carboligation reactions with an exceptionally broad substrate range, a feature that makes KdcA a potentially valuable biocatalyst for C-C bond formation, in particular for the enzymatic synthesis of diversely substituted 2-hydroxyketones with high enantioselectivity
-
-
-
additional information
?
-
P9WG37
enzyme is found to convert a broad spectrum of branched-chain and aromatic alpha-keto acids. Determined the catalytic constants for the conversion of 11 aliphatic, aromatic, and branched-chain alpha-keto acids
-
-
-
additional information
?
-
-
mitochondrial branched-chain aminotransferase binds to the E1 decarboxylase of BCKDC, forming a metabolon that allows channeling of branched-chain alpha-keto acids from mitochondrial branched-chain aminotransferase to E1
-
-
-
additional information
?
-
P9WG37
enzyme is found to convert a broad spectrum of branched-chain and aromatic alpha-keto acids. Determined the catalytic constants for the conversion of 11 aliphatic, aromatic, and branched-chain alpha-keto acids
-
-
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
3-methyl-2-oxopentanoate
CO2 + 2-methylbutanal
show the reaction diagram
-
-
-
-
?
3-methyl-2-oxopentanoate
CO2 + 2-methylbutanal
show the reaction diagram
-
-
-
-
?
additional information
?
-
-
essential for the synthesis of branched-chain fatty acids
-
-
-
additional information
?
-
-
plants with enzymic activity show enhanced cold tolerance, role as a protective mechanism for growth of plants under sub optimal temperatures
-
-
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
thiamine diphosphate
-
stimulates moderately
thiamine diphosphate
-
required
thiamine diphosphate
-
binds to E1b component of enzyme complex
thiamine diphosphate
-
dependent
thiamine diphosphate
P9WG37
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ferricyanide
-
stimulates
Mg2+
-
dependent
additional information
-
no requirement for MgCl2
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
2-[(R)-1-hydroxyethyl]deaza thiamine diphosphate
-
0.1 mM
pyruvate
P9WG37
weak substrate inhibition
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
NaCl
Q6QBS4
20% increase in activity at 1 M
additional information
P9WG37
stopped-flow kinetics show that MtKDC is allosterically activated by 2-oxoacids, also amino acids are potent activators
-
additional information
-
GDH1 alone does not have a significant effect on BCKDC activity. However, addition of GDH1 and glutamate enhance the calculated kcat value by 40-45% with mitochondrial branched-chain aminotransferase present when compared with BCKDC plus branched-chain alpha-keto acids alone
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.09
(indol-3-yl)pyruvate
P9WG37
-
20.25
2-oxo-3-methylpentanoate
P9WG37
-
0.127
2-oxo-3-phenylpropanoate
-
in 50 mM potassium phosphate buffer, pH 6.8 in the presence of 2.5 mM MgSO4 and 0.1 mM thiamine diphosphate
138.3
2-oxobutyrate
P9WG37
-
0.83
2-Oxohexanoate
P9WG37
-
0.6
2-oxohexanoic acid
-
in 50 mM potassium phosphate buffer, pH 6.8 in the presence of 2.5 mM MgSO4 and 0.1 mM thiamine diphosphate
2
2-Oxoisohexanoate
-
25C, pH 7.2
0.05
2-oxoisovalerate
-
mutant D295A, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction; mutant D295A, pH 7.5, 30C, overall reaction; wild-type, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction; wild-type, pH 7.5, 30C, overall reaction
0.14
2-oxoisovalerate
-
mutant Y300A, pH 7.5, 30C, overall reaction
0.225
2-oxoisovalerate
-
mutant R301A, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction
0.252
2-oxoisovalerate
-
mutant Y300F, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction
0.255
2-oxoisovalerate
-
mutant R301A, pH 7.5, 30C, overall reaction
0.337
2-oxoisovalerate
-
mutant Y300A, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction
0.39
2-oxoisovalerate
-
mutant Y300F, pH 7.5, 30C, overall reaction
1.21
2-Oxopentanoate
P9WG37
-
1.3
2-Oxopentanoic acid
-
in 50 mM potassium phosphate buffer, pH 6.8 in the presence of 2.5 mM MgSO4 and 0.1 mM thiamine diphosphate
29.77
2-Oxopropanoic acid
-
in 50 mM potassium phosphate buffer, pH 6.8 in the presence of 2.5 mM MgSO4 and 0.1 mM thiamine diphosphate
0.63
3-(4-hydroxyphenyl)-2-oxopropanoic acid
-
in 50 mM potassium phosphate buffer, pH 6.8 in the presence of 2.5 mM MgSO4 and 0.1 mM thiamine diphosphate
5.02
3-methyl-2-oxobutanoic acid
-
in 50 mM potassium phosphate buffer, pH 6.8 in the presence of 2.5 mM MgSO4 and 0.1 mM thiamine diphosphate
0.87
3-methyl-2-oxopentanoate
-
25C, pH 7.2
1.3
4-(methylsulfanyl)-2-oxobutanoic acid
-
in 50 mM potassium phosphate buffer, pH 6.8 in the presence of 2.5 mM MgSO4 and 0.1 mM thiamine diphosphate
0.264
4-methyl-2-oxopentanoic acid
-
in 50 mM potassium phosphate buffer, pH 6.8 in the presence of 2.5 mM MgSO4 and 0.1 mM thiamine diphosphate
26.11
benzoylformate
P9WG37
-
0.234
Indole-3-pyruvate
-
in 50 mM potassium phosphate buffer, pH 6.8 in the presence of 2.5 mM MgSO4 and 0.1 mM thiamine diphosphate
0.001
L-3-Methyl-2-oxopentanoate
-
and 2-oxoisopentanoate, 2-oxoisohexanoate, value below, 30C, pH 7.5
7.5
oxo(phenyl)acetic acid
-
in 50 mM potassium phosphate buffer, pH 6.8 in the presence of 2.5 mM MgSO4 and 0.1 mM thiamine diphosphate
1.17
phenylpyruvate
P9WG37
-
0.4
pyruvate
P9WG37
-
0.00063
thiamine diphosphate
-
wild-type, pH 7.5, 30C, overall reaction
0.0066
thiamine diphosphate
-
wild-type, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction
0.015
thiamine diphosphate
-
mutant D295A, pH 7.5, 30C, overall reaction
0.016
thiamine diphosphate
-
mutant D295A, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction
0.018
thiamine diphosphate
-
mutant Y300F, pH 7.5, 30C, overall reaction
0.026
thiamine diphosphate
-
mutant R301A, pH 7.5, 30C, overall reaction; mutant Y300A, pH 7.5, 30C, overall reaction
0.028
thiamine diphosphate
-
mutant Y300F, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction
0.029
thiamine diphosphate
-
mutant R301A, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction
0.04
thiamine diphosphate
-
mutant Y300A, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
2.98
(indol-3-yl)pyruvate
P9WG37
-
19.75
2-oxo-3-methylpentanoate
P9WG37
-
5.98
2-oxobutyrate
P9WG37
-
7.06
2-Oxohexanoate
P9WG37
-
31.5
2-Oxoisohexanoate
P9WG37
-
28.35
2-oxoisopentanoate
P9WG37
-
0.19
2-oxoisovalerate
-
wild-type, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction; wild-type, pH 7.5, 30C, overall reaction
0.38
2-oxoisovalerate
-
mutant Y300A, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction; mutant Y300A, pH 7.5, 30C, overall reaction
0.41
2-oxoisovalerate
-
mutant R301A, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction; mutant R301A, pH 7.5, 30C, overall reaction
0.48
2-oxoisovalerate
-
mutant D295A, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction; mutant D295A, pH 7.5, 30C, overall reaction
0.57
2-oxoisovalerate
-
mutant Y300F, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction; mutant Y300F, pH 7.5, 30C, overall reaction
6.95
2-Oxopentanoate
P9WG37
-
29
3-methyl-2-oxobutyrate
-
pH and temperature not specified in the publication
33
3-methyl-2-oxobutyrate
-
pH and temperature not specified in the publication
23
3-methyl-2-oxopentanoate
-
pH and temperature not specified in the publication
25
3-methyl-2-oxopentanoate
-
pH and temperature not specified in the publication
27
3-methyl-2-oxopentanoate
-
pH and temperature not specified in the publication
3.89
4-hydroxyphenylpyruvate
P9WG37
-
7
4-methyl-2-oxopentanoate
-
pH and temperature not specified in the publication
8.5
4-methyl-2-oxopentanoate
-
pH and temperature not specified in the publication
18.57
benzoylformate
P9WG37
-
1.7
phenylpyruvate
-
pH and temperature not specified in the publication
1.8
phenylpyruvate
-
pH and temperature not specified in the publication
7.5
phenylpyruvate
P9WG37
-
2.14
pyruvate
P9WG37
-
0.19
thiamine diphosphate
-
wild-type, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction; wild-type, pH 7.5, 30C, overall reaction
0.31
thiamine diphosphate
-
mutant Y300A, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction; mutant Y300A, pH 7.5, 30C, overall reaction
0.41
thiamine diphosphate
-
mutant R301A, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction; mutant R301A, pH 7.5, 30C, overall reaction
0.42
thiamine diphosphate
-
mutant D295A, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction; mutant D295A, pH 7.5, 30C, overall reaction
0.53
thiamine diphosphate
-
mutant Y300F, pH 7.5, 30C, E1b-catalyzed decarboxylation reaction; mutant Y300F, pH 7.5, 30C, overall reaction
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
124
pyruvate
P9WG37
weak substrate inhibition is detected for pyruvate
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
0.02
-
30C, pH 7.5
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
6.3
Q6QBS4
-
6.5
P9WG37
assay at
7.2
-
assay at
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
6 - 9
-
pH 6: about 60% of maximum activity, pH 9: about 55% of maximum activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
25
-
assay at
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
additional information
-
BCKDC proteins in the metabolon are found in all tissues except skeletal muscle, which harbors only the five subunits
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
59780
P9WG37
calculated from the deduced amino acid sequence
693210
126700
-
calculated from amino acid sequence
677424
146000
-
gel filtration
677424
160000
-
gel filtration
648376
171000
-
with His6-tagged alpha subunits, gel filtration, sucrose density gradient
648372
175000
-
gel filtration
648370
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
?
Q6QBS4
x * 60900, calculated, x * 62000, SDS-PAGE
homodimer
-
x-ray crystallography
tetramer
-
2 * 33000, A-subunit, + 2 * 39800, B-subunit, SDS-PAGE
tetramer
-
2 * 45000, alpha subunit, + 2 * 37000, beta subunit, SDS-PAGE
tetramer
-
2 * 45500, alpha subunit, + 2 * 37800, beta subunit, SDS-PAGE, enzyme forms part of the branched chain alpha-ketoacid dehydrogenase complex
homodimer
-
2 * 73000, gel filtration
additional information
-
assembly of tetramer is promoted by chaperonins groEL and groES
additional information
-
concurrent expression of subunits alpha and beta is important for assembly
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
phosphoprotein
-
when the E1alpha subunit is fully phosphorylated, overall BCKDC activity is zero
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
hanging drop vapour diffusion method using 25-26% (v/v) PEG 200 in 0.1 M MES buffer pH 6.2 as a precipitant
-
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
4
-
rapid inactivation at pH 4
677424
5 - 7
-
stable at pH 5-7
677424
8
-
inactivation at pH 8
677424
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
40
-
sufficiently stable up to 40C, but rapidly loses activity at higher temperatures
677424
ORGANIC SOLVENT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
DMSO
-
completely stable in the presence of 20% (v/v) DMSO (half-life: 150 h)
polyethyleneglycol
-
rapidly inactivated in the presence of 15% (v/v) PEG-400
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
immobilized metal chelate chromatography and gel filtration
-
Ni-NTA column chromatography
-
nickel-charged metal chelating column chromatography
-
anion exchange chromatography and gel filtration
P9WG37
nickel-charged metal chelating column chromatography
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli
-
expressed in Escherichia coli BL21 cells
-
expressed in Escherichia coli strain BL21(DE3)
-
expressed in Escherichia coli strain H-81
-
expressed in Escherichia coli Rosetta 2 (DE3) and various Escherichia coli strains
P9WG37
expressed in Escherichia coli BL21 cells
-
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
A240P-alpha
-
slow assembly of alpha2beta2 tetramer
D295A
-
of component E1b, 2fold increase in kcat-value
E193A-alpha
-
complete inactivation of enzyme
E76A-beta
-
complete inactivation of enzyme
F364C-alpha
-
slow assembly of enzyme, alphabeta dimer
G204S-alpha
-
slow assembly of alpha2beta2 tetramer
G245R-alpha
-
moderately slow assembly of alpha2beta2 tetramer
H146A-beta
-
complete inactivation of enzyme
H291A-alpha
-
partially active enzyme
N222S-alpha
-
markedly higher Km value for thiamin diphosphate
N222S-alpha
-
normal assembly of alpha2beta2 tetramer
R114W-alpha
-
naturally occuring mutation in maple syrup disease patients, strongly reduced binding of thiamin diphosphate, enzyme inactive
R220W-alpha
-
normal assembly of alpha2beta2 tetramer
R220W-alpha
-
naturally occuring mutation in maple syrup disease patients, strongly reduced binding of thiamin diphosphate, enzyme inactive
R287A
-
of component E1b, 40fold increase in Km-value for 2-oxoisovalerate
R301A
-
of component E1b, 4fold increase in Km-value for 2-oxoisovalerate
T166M-alpha
-
enzyme inactive, with attenuated ability to bind thiamin diphosphate
T265R
-
no assembly of alpha2beta2 tetramer
Y224A-alpha
-
markedly higher Km value for thiamin diphosphate
Y300A
-
of component E1b, 6fold increase in Km-value for 2-oxoisovalerate
Y300F
-
of component E1b, 5fold increase in Km-value for 2-oxoisovalerate
Y368C-alpha
-
slow assembly of enzyme as alphabeta dimer and alpha2beta2 tetramer
Y393N-alpha
-
slow assembly of enzyme, alphabeta dimer
Y393N-alpha
-
no assembly of heterotetramer
Renatured/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
after denaturation in 8 M urea, reconstitution at 23C with absolute requirement of chaperonins GroEL/GroES and MgATP2-
-
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
agriculture
-
plants with enzymic activity show enhanced cold tolerance, role as a protective mechanism for growth of plants under sub optimal temperatures
medicine
-
in patients with maple syrup urine disease, mutations such as R114W-alpha or R220W-alpha cause a strongly reduced binding of thiamin diphosphate, rendering the enzyme inactive
medicine
-
naturally occuring mutations cause an impaired assembly of enzyme in patients with maple syrup urine disease
synthesis
-
the enzyme is able to catalyze carboligation reactions with an exceptionally broad substrate range, a feature that makes KdcA a potentially valuable biocatalyst for C-C bond formation, in particular for the enzymatic synthesis of diversely substituted 2-hydroxyketones with high enantioselectivity