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Literature summary extracted from

  • Tyagi, R.; Lee, Y.T.; Guddat, L.W.; Duggleby, R.G.
    Probing the mechanism of the bifunctional enzyme ketol-acid reductoisomerase by site-directed mutagenesis of the active site (2005), FEBS J., 272, 593-602.
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.1.1.86
-
Escherichia coli

Protein Variants

EC Number Protein Variants Comment Organism
1.1.1.86 D217E less than 4% reductoisomerase activity in comparison to wild-type enzyme Escherichia coli
1.1.1.86 D217N less than 4% reductoisomerase activity in comparison to wild-type enzyme Escherichia coli
1.1.1.86 E213D 75% reductoisomerase activity in comparison of wild-type enzyme Escherichia coli
1.1.1.86 E213Q less than 4% reductoisomerase activity in comparison of wild-type enzyme Escherichia coli
1.1.1.86 E221D less than 4% reductoisomerase activity in comparison to wild-type enzyme Escherichia coli
1.1.1.86 E221Q less than 4% reductoisomerase activity in comparison to wild-type enzyme Escherichia coli
1.1.1.86 E389D less than 4% reductoisomerase activity in comparison to wild-type enzyme Escherichia coli
1.1.1.86 E389Q less than 4% reductoisomerase activity in comparison to wild-type enzyme Escherichia coli
1.1.1.86 E393D less than 4% reductoisomerase activity in comparison to wild-type enzyme Escherichia coli
1.1.1.86 E393Q the mutant is insoluble, a soluble form is obtained only after denaturation Escherichia coli
1.1.1.86 H132K less than 4% reductoisomerase activity in comparison of wild-type enzyme Escherichia coli
1.1.1.86 H132Q less than 4% reductoisomerase activity in comparison of wild-type enzyme Escherichia coli
1.1.1.86 K155E less than 4% reductoisomerase activity in comparison of wild-type enzyme Escherichia coli
1.1.1.86 K155Q less than 4% reductoisomerase activity in comparison of wild-type enzyme Escherichia coli
1.1.1.86 K155R less than 4% reductoisomerase activity in comparison of wild-type enzyme Escherichia coli
1.1.1.86 S414A less than 4% reductoisomerase activity in comparison to wild-type enzyme Escherichia coli
1.1.1.86 S414T less than 4% reductoisomerase activity in comparison to wild-type enzyme Escherichia coli

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.1.1.86 0.00016
-
NADPH pH 8.0, 37°C, mutant E213D Escherichia coli
1.1.1.86 0.00253
-
NADPH pH 8.0, 37°C, wild-type enzyme Escherichia coli
1.1.1.86 0.0031
-
NADPH pH 8.0, 37°C, mutant H132K Escherichia coli
1.1.1.86 0.0048
-
NADPH pH 8.0, 37°C, mutant E393D Escherichia coli
1.1.1.86 0.005
-
NADPH pH 8.0, 37°C, mutant D217N Escherichia coli
1.1.1.86 0.005
-
NADPH pH 8.0, 37°C, mutant S414T Escherichia coli
1.1.1.86 0.0073
-
NADPH pH 8.0, 37°C, mutant K155R Escherichia coli
1.1.1.86 0.00804
-
NADPH pH 8.0, 37°C, mutant K155E Escherichia coli
1.1.1.86 0.0084
-
NADPH pH 8.0, 37°C, mutant S414A Escherichia coli
1.1.1.86 0.0093
-
NADPH pH 8.0, 37°C, mutant K155Q Escherichia coli
1.1.1.86 0.02
-
NADPH pH 8.0, 37°C, mutant E221D Escherichia coli
1.1.1.86 0.02
-
NADPH pH 8.0, 37°C, mutant E221Q Escherichia coli
1.1.1.86 0.023
-
NADPH pH 8.0, 37°C, mutant E389D Escherichia coli
1.1.1.86 0.069
-
NADPH pH 8.0, 37°C, mutant H132Q Escherichia coli
1.1.1.86 0.08
-
NADPH pH 8.0, 37°C, mutant D217E Escherichia coli
1.1.1.86 0.17
-
2-ketopantoate pH 8.0, 37°C, wild-type enzyme Escherichia coli
1.1.1.86 0.21
-
3-hydroxy-2-ketobutyrate pH 8.0, 37°C, wild-type enzyme Escherichia coli
1.1.1.86 0.25
-
2-acetolactate pH 8.0, 37°C, wild-type enzyme Escherichia coli
1.1.1.86 0.27
-
3-hydroxy-3-methyl-2-ketobutyrate pH 8.0, 37°C, wild-type enzyme Escherichia coli
1.1.1.86 0.334
-
3-hydroxypyruvate pH 8.0, 37°C, mutant S414A Escherichia coli
1.1.1.86 0.356
-
2-acetolactate pH 8.0, 37°C, mutant E221D Escherichia coli
1.1.1.86 0.414
-
2-acetolactate pH 8.0, 37°C, mutant S414T Escherichia coli
1.1.1.86 0.441
-
3-hydroxypyruvate pH 8.0, 37°C, mutant E213Q Escherichia coli
1.1.1.86 0.588
-
3-hydroxypyruvate pH 8.0, 37°C, mutant E393Q Escherichia coli
1.1.1.86 0.711
-
2-acetolactate pH 8.0, 37°C, mutant S414A Escherichia coli
1.1.1.86 0.818
-
3-hydroxypyruvate pH 8.0, 37°C, mutant H132K Escherichia coli
1.1.1.86 0.922
-
2-acetolactate pH 8.0, 37°C, mutant H213D Escherichia coli
1.1.1.86 0.929
-
2-acetolactate pH 8.0, 37°C, mutant H132Q Escherichia coli
1.1.1.86 1.101
-
3-hydroxypyruvate pH 8.0, 37°C, mutant S414T Escherichia coli
1.1.1.86 1.218
-
2-acetolactate pH 8.0, 37°C, mutant H155R Escherichia coli
1.1.1.86 1.37
-
3-hydroxypyruvate pH 8.0, 37°C, mutant E221D Escherichia coli
1.1.1.86 1.54
-
pyruvate pH 8.0, 37°C, wild-type enzyme Escherichia coli
1.1.1.86 2.028
-
2-acetolactate pH 8.0, 37°C, mutant E289D Escherichia coli
1.1.1.86 2.66
-
3-hydroxypyruvate pH 8.0, 37°C, mutant K155E Escherichia coli
1.1.1.86 2.96
-
3-hydroxypyruvate pH 8.0, 37°C, wild-type enzyme Escherichia coli
1.1.1.86 3.15
-
2-ketovalerate pH 8.0, 37°C, wild-type enzyme Escherichia coli
1.1.1.86 3.32
-
3-hydroxypyruvate pH 8.0, 37°C, mutant E393D Escherichia coli
1.1.1.86 3.67
-
3-hydroxypyruvate pH 8.0, 37°C, mutant E213D Escherichia coli
1.1.1.86 4.56
-
2-Ketobutyrate pH 8.0, 37°C, wild-type enzyme Escherichia coli
1.1.1.86 6.91
-
2-ketoisovalerate pH 8.0, 37°C, wild-type enzyme Escherichia coli
1.1.1.86 7.43
-
3-hydroxypyruvate pH 8.0, 37°C, mutant H132Q Escherichia coli
1.1.1.86 7.64
-
3-hydroxypyruvate pH 8.0, 37°C, mutant D217N Escherichia coli
1.1.1.86 8.5
-
3-hydroxypyruvate pH 8.0, 37°C, mutant E389D Escherichia coli
1.1.1.86 8.88
-
3-hydroxypyruvate pH 8.0, 37°C, mutant E389Q Escherichia coli
1.1.1.86 13.6
-
3-hydroxypyruvate pH 8.0, 37°C, mutant K155R Escherichia coli
1.1.1.86 15.3
-
3-hydroxypyruvate pH 8.0, 37°C, mutant K155Q Escherichia coli

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
1.1.1.86 Co2+ the reaction requires a divalent metal ion Escherichia coli
1.1.1.86 Mg2+ the reaction requires a divalent metal ion Escherichia coli
1.1.1.86 Mn2+ the reaction requires a divalent metal ion Escherichia coli

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.1.1.86 additional information Escherichia coli the enzyme is involved in the biosynthesis of the branched-chain amino acids ?
-
?

Organism

EC Number Organism UniProt Comment Textmining
1.1.1.86 Escherichia coli
-
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.1.1.86 of wild-type and mutants Escherichia coli

Reaction

EC Number Reaction Comment Organism Reaction ID
1.1.1.86 (2R)-2,3-dihydroxy-3-methylbutanoate + NADP+ = (2S)-2-hydroxy-2-methyl-3-oxobutanoate + NADPH + H+ bifunctional enzyme that catalyses two different reactions at a common active site, an isomerization consisting of an alkyl migration, followed by an NADPH-dependent reduction of a-ketoacid Escherichia coli

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
1.1.1.86 additional information
-
the specific activity with pyruvate is 1% and with 2-ketovalerate, 2-ketopantoate and 2-ketobutyrate is 8% of that of 2-acetolactate, comparison of activities of wild-type and mutant enzymes Escherichia coli

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.1.1.86 2-acetolactate + NADPH + H+
-
Escherichia coli 2,3-dihydroxy-3-methylbutanoate + NADP+
-
r
1.1.1.86 2-ketobutyrate + NADPH + H+
-
Escherichia coli ?
-
r
1.1.1.86 2-ketoisovalerate + NADPH + H+
-
Escherichia coli ?
-
r
1.1.1.86 2-ketopantoate + NADPH + H+
-
Escherichia coli ?
-
r
1.1.1.86 2-ketovalerate + NADPH + H+
-
Escherichia coli ?
-
r
1.1.1.86 3-hydroxy-2-ketobutyrate + NADPH + H+
-
Escherichia coli ?
-
r
1.1.1.86 3-hydroxy-3-methyl-2-ketobutyrate + NADP+
-
Escherichia coli ?
-
r
1.1.1.86 3-hydroxypyruvate + NADPH + H+
-
Escherichia coli ?
-
r
1.1.1.86 additional information the enzyme is involved in the biosynthesis of the branched-chain amino acids Escherichia coli ?
-
?
1.1.1.86 pyruvate + NADPH + H+
-
Escherichia coli ?
-
r

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.1.1.86 0.021
-
pyruvate pH 8.0, 37°C Escherichia coli
1.1.1.86 0.05
-
2-ketovalerate pH 8.0, 37°C Escherichia coli
1.1.1.86 0.167
-
2-Ketobutyrate pH 8.0, 37°C Escherichia coli
1.1.1.86 0.182
-
2-ketoisovalerate pH 8.0, 37°C Escherichia coli
1.1.1.86 0.194
-
2-ketopantoate pH 8.0, 37°C Escherichia coli
1.1.1.86 0.594
-
3-hydroxy-2-ketobutyrate pH 8.0, 37°C Escherichia coli
1.1.1.86 2.231
-
2-acetolactate pH 8.0, 37°C Escherichia coli
1.1.1.86 3.511
-
3-hydroxy-3-methyl-2-ketobutyrate pH 8.0, 37°C Escherichia coli
1.1.1.86 5.376
-
3-hydroxypyruvate pH 8.0, 37°C Escherichia coli

Cofactor

EC Number Cofactor Comment Organism Structure
1.1.1.86 NADP+
-
Escherichia coli
1.1.1.86 NADPH
-
Escherichia coli