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Literature summary for 4.1.1.7 extracted from

  • Lingen, B.; Grotzinger, J.; Kolter, D.; Kula, M.R.; Pohl, M.
    Improving the carboligase activity of benzoylformate decarboxylase from Pseudomonas putida by a combination of directed evolution and site-directed mutagenesis (2002), Protein Eng., 15, 585-593.
    View publication on PubMed
Search for more literature for 4.1.1.7

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli strain SG13009/pREP4 Pseudomonas putida

Protein Variants

Protein Variants Comment Organism
L476A has a 4.11fold higher carboligase activity than the wild type enzyme Pseudomonas putida
L476C has a 4.31fold higher carboligase activity than the wild type enzyme Pseudomonas putida
L476G has a 4.27fold higher carboligase activity than the wild type enzyme Pseudomonas putida
L476H has a 3.19fold higher carboligase activity than the wild type enzyme Pseudomonas putida
L476K has a 5fold higher carboligase activity than the wild type enzyme Pseudomonas putida
L476M has a 4.23fold higher carboligase activity than the wild type enzyme Pseudomonas putida
L476P decreased activity Pseudomonas putida
L476Q has a 5.06fold higher carboligase activity than the wild type enzyme Pseudomonas putida
L476Q/S535G decreased activity Pseudomonas putida
L476S has a 3.5fold higher carboligase activity than the wild type enzyme Pseudomonas putida
L476T has a 3.98fold higher carboligase activity than the wild type enzyme Pseudomonas putida
S181T has a 1.75fold higher carboligase activity than the wild type enzyme Pseudomonas putida
S181T/L476P increased activity Pseudomonas putida

General Stability

General Stability Organism
the wild type enzyme shows no loss of activity in cofactor-free buffer over 24 h Pseudomonas putida

Inhibitors

Inhibitors Comment Organism Structure
benzoylformate
-
 Pseudomonas putida

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.54
-
 benzoylformate wild type enzyme, in 50 mM potassium phosphate buffer pH 7.0, 0.5 mM thiamine diphosphate, 2.5 mM MgSO4, at 30°C Pseudomonas putida
0.58
-
 benzoylformate mutant enzyme L476Q, in 50 mM potassium phosphate buffer pH 7.0, 0.5 mM thiamine diphosphate, 2.5 mM MgSO4, at 30°C Pseudomonas putida
0.59
-
 benzoylformate mutant enzyme L476G, in 50 mM potassium phosphate buffer pH 7.0, 0.5 mM thiamine diphosphate, 2.5 mM MgSO4, at 30°C Pseudomonas putida
0.6
-
 benzoylformate mutant enzyme S181T/L476P, in 50 mM potassium phosphate buffer pH 7.0, 0.5 mM thiamine diphosphate, 2.5 mM MgSO4, at 30°C Pseudomonas putida
1
-
 benzoylformate mutant enzyme L476H, in 50 mM potassium phosphate buffer pH 7.0, 0.5 mM thiamine diphosphate, 2.5 mM MgSO4, at 30°C Pseudomonas putida
1.02
-
 benzoylformate mutant enzyme L476T, in 50 mM potassium phosphate buffer pH 7.0, 0.5 mM thiamine diphosphate, 2.5 mM MgSO4, at 30°C Pseudomonas putida
1.03
-
 benzoylformate mutant enzyme L476P, in 50 mM potassium phosphate buffer pH 7.0, 0.5 mM thiamine diphosphate, 2.5 mM MgSO4, at 30°C Pseudomonas putida
1.06
-
 benzoylformate mutant enzyme L476Q/S525G, in 50 mM potassium phosphate buffer pH 7.0, 0.5 mM thiamine diphosphate, 2.5 mM MgSO4, at 30°C Pseudomonas putida
1.08
-
 benzoylformate mutant enzyme L476M, in 50 mM potassium phosphate buffer pH 7.0, 0.5 mM thiamine diphosphate, 2.5 mM MgSO4, at 30°C Pseudomonas putida
1.14
-
 benzoylformate mutant enzyme L476S, in 50 mM potassium phosphate buffer pH 7.0, 0.5 mM thiamine diphosphate, 2.5 mM MgSO4, at 30°C Pseudomonas putida
1.17
-
 benzoylformate mutant enzyme L476A, in 50 mM potassium phosphate buffer pH 7.0, 0.5 mM thiamine diphosphate, 2.5 mM MgSO4, at 30°C Pseudomonas putida
1.23
-
 benzoylformate mutant enzyme S181T, in 50 mM potassium phosphate buffer pH 7.0, 0.5 mM thiamine diphosphate, 2.5 mM MgSO4, at 30°C Pseudomonas putida
1.26
-
 benzoylformate mutant enzyme L476C, in 50 mM potassium phosphate buffer pH 7.0, 0.5 mM thiamine diphosphate, 2.5 mM MgSO4, at 30°C Pseudomonas putida
1.46
-
 benzoylformate mutant enzyme L476K, in 50 mM potassium phosphate buffer pH 7.0, 0.5 mM thiamine diphosphate, 2.5 mM MgSO4, at 30°C Pseudomonas putida

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required Pseudomonas putida

Organic Solvent Stability

Organic Solvent Comment Organism
DMSO retains its full activity after 10 days of incubation in DMSO (20% v/v) at 25°C Pseudomonas putida
Ethanol retains its full activity after 10 days of incubation in aqueous ethanol (1.5 M) at 25°C Pseudomonas putida

Organism

Organism UniProt Comment Textmining
Pseudomonas putida
-
-
-

Purification (Commentary)

Purification (Comment) Organism
Ni-NTA column chromatography Pseudomonas putida

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
benzaldehyde + acetaldehyde
-
 Pseudomonas putida (S)-2-hydroxy-1-phenyl-propanone
-
 ?
benzoylformate
-
 Pseudomonas putida benzaldehyde + CO2
-
 ?

Synonyms

Synonyms Comment Organism
BFD
-
 Pseudomonas putida

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
8
-
 the optimal pH is 8.0 or above for the carboligase reaction Pseudomonas putida

Cofactor

Cofactor Comment Organism Structure
thiamine diphosphate dependent Pseudomonas putida

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
45
-
 benzoylformate mutant enzyme L476P Pseudomonas putida
143
-
 benzoylformate wild type enzyme Pseudomonas putida