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Literature summary for 4.1.1.47 extracted from

  • Nemeria, N.; Binshtein, E.; Patel, H.; Balakrishnan, A.; Vered, I.; Shaanan, B.; Barak, Z.; Chipman, D.; Jordan, F.
    Glyoxylate carboligase: a unique thiamin diphosphate-dependent enzyme that can cycle between the 4'-aminopyrimidinium and 1,4'-iminopyrimidine tautomeric forms in the absence of the conserved glutamate (2012), Biochemistry, 51, 7940-7952.
    View publication on PubMedView publication on EuropePMC

Protein Variants

Protein Variants Comment Organism
I393A the mutation leads to a lower catalytic efficiency (3.9%) compared to the wild type enzyme. The enzyme is converted to an acetolactate synthase which can use pyruvate as a substrate with a catalytic efficiency (kcat/Km) of about 20times higher than that of the wild type enzyme Escherichia coli
I393V the mutation leads to a lower catalytic efficiency (5.3%) compared to the wild type enzyme Escherichia coli
I479V the mutation leads to a lower catalytic efficiency (4.8%) compared to the wild type enzyme Escherichia coli
L478A the mutation leads to a lower catalytic efficiency (0.34%) compared to the wild type enzyme Escherichia coli
V51D the substitution shifts the pH optimum to 6.0-6.2, the mutant is less active (1.2%) than the wild type enzyme (turnover rates are 2 orders of magnitude lower) despite having higher rate of activation of the coenzyme Escherichia coli
V51D/I393A the enzyme is converted to an acetolactate synthase which can use pyruvate as a substrate with a catalytic efficiency (kcat/Km) of about 20times higher than that of the wild type enzyme Escherichia coli
V51E the mutant is less active than the wild type enzyme (turnover rates are 7fold lower) despite having higher rate of activation of the coenzyme Escherichia coli

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
Glyoxylate Escherichia coli
-
Tartronate semialdehyde + CO2
-
?

Organism

Organism UniProt Comment Textmining
Escherichia coli
-
-
-

Purification (Commentary)

Purification (Comment) Organism
HiTrap Chelating Ni column chromatography Escherichia coli

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
0.06
-
mutant enzyme L478A, in 50 mM KH2PO4 (pH 7.7), 0.06 M KCl, 0.1 mM thiamin diphosphate, 5 mM MgCl2, at 37°C Escherichia coli
0.2
-
mutant enzyme V51D, in 50 mM KH2PO4 (pH 7.7), 0.06 M KCl, 0.1 mM thiamin diphosphate, 5 mM MgCl2, at 37°C Escherichia coli
0.68
-
mutant enzyme I393A, in 50 mM KH2PO4 (pH 7.7), 0.06 M KCl, 0.1 mM thiamin diphosphate, 5 mM MgCl2, at 37°C Escherichia coli
0.84
-
mutant enzyme I479V, in 50 mM KH2PO4 (pH 7.7), 0.06 M KCl, 0.1 mM thiamin diphosphate, 5 mM MgCl2, at 37°C Escherichia coli
0.93
-
mutant enzyme I393V, in 50 mM KH2PO4 (pH 7.7), 0.06 M KCl, 0.1 mM thiamin diphosphate, 5 mM MgCl2, at 37°C Escherichia coli
17.5
-
wild type enzyme, in 50 mM KH2PO4 (pH 7.7), 0.06 M KCl, 0.1 mM thiamin diphosphate, 5 mM MgCl2, at 37°C Escherichia coli

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
Glyoxylate
-
Escherichia coli Tartronate semialdehyde + CO2
-
?
additional information the enzyme is unreactive with 2-ketoacids Escherichia coli ?
-
?

Synonyms

Synonyms Comment Organism
GCL
-
Escherichia coli
Glyoxylate carboligase
-
Escherichia coli

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7 7.7
-
Escherichia coli

Cofactor

Cofactor Comment Organism Structure
thiamine diphosphate dependent on Escherichia coli

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.58
-
glyoxylate mutant enzyme I479V, in 50 mM KH2PO4 (pH 7.7), 0.06 M KCl, 0.1 mM thiamin diphosphate, 5 mM MgCl2, at 37°C Escherichia coli
0.65
-
glyoxylate mutant enzyme I393V, in 50 mM KH2PO4 (pH 7.7), 0.06 M KCl, 0.1 mM thiamin diphosphate, 5 mM MgCl2, at 37°C Escherichia coli
1.3
-
glyoxylate mutant enzyme V51D, in 50 mM KH2PO4 (pH 7.7), 0.06 M KCl, 0.1 mM thiamin diphosphate, 5 mM MgCl2, at 37°C Escherichia coli
1.79
-
glyoxylate mutant enzyme L478A, in 50 mM KH2PO4 (pH 7.7), 0.06 M KCl, 0.1 mM thiamin diphosphate, 5 mM MgCl2, at 37°C Escherichia coli
2.06
-
glyoxylate mutant enzyme I393A, in 50 mM KH2PO4 (pH 7.7), 0.06 M KCl, 0.1 mM thiamin diphosphate, 5 mM MgCl2, at 37°C Escherichia coli
21
-
glyoxylate wild type enzyme, in 50 mM KH2PO4 (pH 7.7), 0.06 M KCl, 0.1 mM thiamin diphosphate, 5 mM MgCl2, at 37°C Escherichia coli