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Literature summary for 4.1.1.7 extracted from

  • Yep, A.; McLeish, M.J.
    Engineering the substrate binding site of benzoylformate decarboxylase (2009), Biochemistry, 48, 8387-8395.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli BL21(DE3)pLysS cells Pseudomonas putida

Protein Variants

Protein Variants Comment Organism
A460Y the mutant shows significantly improved catalytic activity toward pyruvate Pseudomonas putida
T377L the mutant shows significantly improved catalytic activity toward pyruvate Pseudomonas putida
T377L/A460Y the double mutant shows significantly improved catalytic activity toward pyruvate (11000fold improvement in the ratio between pyruvate and benzoylformate utilization) Pseudomonas putida

Inhibitors

Inhibitors Comment Organism Structure
(R)-mandelate substrate analogue inhibitor Pseudomonas putida

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.04
-
benzylformate mutant enzyme A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
0.1
-
2-ketopentanoate mutant enzyme T377L/A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
0.15
-
2-ketohexanoate mutant enzyme T377L/A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
0.27
-
benzylformate wild type enzyme, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
0.28
-
2-ketopentanoate mutant enzyme A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
0.28
-
2-ketohexanoate mutant enzyme A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
0.42
-
2-ketobutanoate mutant enzyme T377L/A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
0.45
-
benzylformate mutant enzyme T377L, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
1.1
-
2-ketopentanoate mutant enzyme T377L, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
1.3
-
2-ketobutanoate mutant enzyme A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
1.3
-
2-ketohexanoate mutant enzyme T377L, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
1.5
-
benzylformate mutant enzyme T377L/A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
2
-
pyruvate mutant enzyme T377L/A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
4.1
-
2-ketohexanoate wild type enzyme, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
4.3
-
2-ketobutanoate mutant enzyme T377L, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
4.8
-
pyruvate mutant enzyme T377L, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
6
-
2-ketopentanoate wild type enzyme, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
7.5
-
2-ketobutanoate wild type enzyme, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
21
-
pyruvate mutant enzyme A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida

Organism

Organism UniProt Comment Textmining
Pseudomonas putida P20906
-
-

Purification (Commentary)

Purification (Comment) Organism
-
Pseudomonas putida

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2-ketohexanoate
-
Pseudomonas putida valeraldehyde + CO2
-
?
2-ketopentanoate
-
Pseudomonas putida butyraldehyde + CO2
-
?
2-oxobutanoate
-
Pseudomonas putida propionaldehyde + CO2
-
?
benzylformate
-
Pseudomonas putida benzaldehyde + CO2
-
?
pyruvate pyruvate is no substrate for the wild type enzyme Pseudomonas putida acetaldehyde + CO2
-
?

Synonyms

Synonyms Comment Organism
BFDC
-
Pseudomonas putida

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.81
-
2-ketopentanoate mutant enzyme T377L/A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
1.1
-
2-ketohexanoate mutant enzyme T377L, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
1.2
-
2-ketohexanoate mutant enzyme T377L/A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
1.4
-
pyruvate mutant enzyme A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
1.7
-
benzylformate mutant enzyme A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
2
-
pyruvate mutant enzyme T377L, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
3.3
-
2-ketopentanoate mutant enzyme T377L, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
3.6
-
pyruvate mutant enzyme T377L/A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
4
-
2-ketobutanoate wild type enzyme, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
4.4
-
2-ketohexanoate mutant enzyme A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
5.3
-
2-ketohexanoate wild type enzyme, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
5.8
-
2-ketobutanoate mutant enzyme T377L/A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
6
-
2-ketobutanoate mutant enzyme A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
6.2
-
2-ketopentanoate mutant enzyme A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
7.5
-
2-ketobutanoate mutant enzyme T377L, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
11
-
2-ketopentanoate wild type enzyme, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
25
-
benzylformate mutant enzyme T377L/A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
110
-
benzylformate mutant enzyme T377L, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
320
-
benzylformate wild type enzyme, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida

Cofactor

Cofactor Comment Organism Structure
thiamine diphosphate
-
Pseudomonas putida

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.012
-
pyruvate wild type enzyme, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
0.065
-
pyruvate mutant enzyme A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
0.42
-
pyruvate mutant enzyme T377L, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
0.5
-
2-ketobutanoate wild type enzyme, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
0.86
-
2-ketohexanoate mutant enzyme T377L, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
1.3
-
2-ketohexanoate wild type enzyme, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
1.7
-
2-ketobutanoate mutant enzyme T377L, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
1.8
-
pyruvate mutant enzyme T377L/A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
1.8
-
2-ketopentanoate wild type enzyme, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
3
-
2-ketopentanoate mutant enzyme T377L, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
4.6
-
2-ketobutanoate mutant enzyme A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
8
-
2-ketohexanoate mutant enzyme T377L/A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
8.1
-
2-ketopentanoate mutant enzyme T377L/A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
14
-
2-ketobutanoate mutant enzyme T377L/A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
16
-
2-ketohexanoate mutant enzyme A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
16
-
benzylformate mutant enzyme T377L/A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
22
-
2-ketopentanoate mutant enzyme A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
43
-
benzylformate mutant enzyme A460Y, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
244
-
benzylformate mutant enzyme T377L, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida
1180
-
benzylformate wild type enzyme, in 100 mM potassium phosphate buffer (pH 6.0), at 30°C Pseudomonas putida