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Literature summary for 5.4.4.2 extracted from

  • Sridharan, S.; Howard, N.; Kerbarh, O.; B?aszczyk, M.; Abell, C.; Blundell, T.L.
    Crystal structure of Escherichia coli enterobactin-specific isochorismate synthase (EntC) bound to its reaction product isochorismate: implications for the enzyme mechanism and differential activity of chorismate-utilizing enzymes (2010), J. Mol. Biol., 397, 290-300.
    View publication on PubMed

Cloned(Commentary)

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

Crystallization (Commentary)

Crystallization (Comment) Organism
in complex with isochorismate and Mg2+, hanging drop vapor diffusion method, using 100 mM MES buffer pH 6.5, 12% PEG (w/v) 20000 Escherichia coli

Protein Variants

Protein Variants Comment Organism
A303T the kcat/Km (chorismate) for the A303T mutant is 130fold lower than that for wild type enzyme Escherichia coli
A303T/F327Y the mutant shows no detectable activity Escherichia coli
A303T/F327Y/F359Q the mutant shows no detectable activity Escherichia coli
A303T/F327Y/F359Q/I346L the mutant shows no detectable activity Escherichia coli
A303T/F327Y/I346L the mutant shows no detectable activity Escherichia coli
A303T/F359Q the mutant shows no detectable activity Escherichia coli
F327Y the mutant displays reduced activity compared to the wild type enzyme Escherichia coli
F327Y/F359Q the mutant displays a reduced kcat/Km value compared to the wild type enzyme Escherichia coli
F327Y/F359Q/I346L the mutant shows no detectable activity Escherichia coli
F359Q the mutant displays a reduced kcat/Km value compared to the wild type enzyme Escherichia coli
F359Q the mutation causes loss of catalytic activity Escherichia coli
F359Q/I346L the mutant displays a reduced kcat/Km value compared to the wild type enzyme Escherichia coli
I346L the EntC mutant has a 12fold lower kcat/Km (chorismate) than the wild type enzyme Escherichia coli
L304A the mutation causes loss of catalytic activity Escherichia coli
L304A/F359Q the mutant shows no detectable activity Escherichia coli

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.007
-
chorismate wild type enzyme, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.04
-
chorismate mutant enzyme F327Y/F359Q, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.066
-
chorismate mutant enzyme F359Q, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.076
-
chorismate mutant enzyme F327Y, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.076
-
chorismate mutant enzyme I346L, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.103
-
chorismate mutant enzyme A303T, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.261
-
chorismate mutant enzyme F359Q/I346L, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.466
-
chorismate mutant enzyme F327Y/I346L, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ the enzyme contains Mg2+ Escherichia coli

Organism

Organism UniProt Comment Textmining
Escherichia coli P0AEJ2
-
-

Purification (Commentary)

Purification (Comment) Organism
Ni2+-charged HiTrap chelating agarose column chromatography and Superdex 75 gel filtration Escherichia coli

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
Chorismate
-
Escherichia coli Isochorismate
-
?

Synonyms

Synonyms Comment Organism
EntC
-
Escherichia coli
ICS
-
Escherichia coli

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.05
-
chorismate mutant enzyme F327Y/I346L, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.067
-
chorismate mutant enzyme A303T, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.13
-
chorismate mutant enzyme F327Y, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.18
-
chorismate mutant enzyme F359Q/I346L, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.43
-
chorismate mutant enzyme F327Y/F359Q, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.55
-
chorismate mutant enzyme I346L, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.6
-
chorismate mutant enzyme F359Q, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.67
-
chorismate wild type enzyme, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.11
-
chorismate mutant enzyme F327Y/I346L, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.65
-
chorismate mutant enzyme A303T, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
0.69
-
chorismate mutant enzyme F359Q/I346L, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
1.71
-
chorismate mutant enzyme F327Y, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
7.24
-
chorismate mutant enzyme I346L, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
9.1
-
chorismate mutant enzyme F359Q, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
10.75
-
chorismate mutant enzyme F327Y/F359Q, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli
95.7
-
chorismate wild type enzyme, in 100 mM phosphate pH 7.0, 10 mM MgCl2 Escherichia coli