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Literature summary for 6.3.2.14 extracted from

  • Sikora, A.L.; Wilson, D.J.; Aldrich, C.C.; Blanchard, J.S.
    Kinetic and inhibition studies of dihydroxybenzoate-AMP ligase from Escherichia coli (2010), Biochemistry, 49, 3648-3657.
    View publication on PubMedView publication on EuropePMC
Search for more literature for 6.3.2.14

Cloned(Commentary)

Cloned (Comment) Organism
gene entE, expression in Escherichia coli strain BL21(DE3) Escherichia coli
gene entE, expression of His-tagged EntE in Escherichia coli strain BL21(DE3) Escherichia coli

Inhibitors

Inhibitors Comment Organism Structure
5'-O-[N-(2,3-dihydroxybenzoyl)sulfamoyl]adenosine
-
 Escherichia coli
5'-O-[N-(salicyl)sulfamoyl]adenosine
-
 Escherichia coli

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information two possible kinetic mechanisms can explain nonlinear kinetics: one-step slow association and two-step isomerization, bi-uni-uni-bi ping-pong kinetic mechanism, kinetic analysis, overview Escherichia coli
0.0025
-
 2,3-Dihydroxybenzoate pH 7.8, 25°C, recombinant EntE Escherichia coli
0.0029
-
 phosphopantetheinylated EntB pH 7.8, 25°C, recombinant EntE Escherichia coli
0.07
-
 salicylic acid pH 7.8, 25°C, recombinant EntE Escherichia coli
0.07
-
 3-hydroxybenzoate pH 7.8, 25°C, recombinant EntE Escherichia coli
0.43
-
 ATP pH 7.8, 25°C, recombinant EntE Escherichia coli
3.1
-
 4-aminosalicylic acid pH 7.8, 25°C, recombinant EntE Escherichia coli

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required Escherichia coli

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
6 ATP + 3 2,3-dihydroxybenzoate + 3 L-serine Escherichia coli overall reaction. EntB, -D, -E, and -F are then required to catalyze the ATP-dependent assembly of enterobactin from three molecules each of 2,3-dihydroxybenzoate and L-serine. EntD, a phosphopantetheinyl transferase, uses coenzyme A to phosphopantetheinylate S245 of the aryl carrier protein domain of EntB. Next, EntE catalyzes the transfer of 2,3-dihydroxybenzoate onto the phosphopantetheinylated (holo) EntB to yield the covalently arylated EntB. Finally, arylated EntB, ATP, and L-serine are used as substrates for the reaction catalyzed by EntF to generate enterobactin enterobactin + 6 AMP + 6 diphosphate
-
 ?
6 ATP + 3 2,3-dihydroxybenzoate + 3 L-serine Escherichia coli overall reaction. EntB, -D, -E, and -F are then required to catalyze the ATP-dependent assembly of enterobactin from three molecules each of 2,3-dihydroxybenzoate and L-serine. EntD, a phosphopantetheinyl transferase, uses coenzyme A to phosphopantetheinylate S245 of the aryl carrier protein domain of EntB. Next, EntE catalyzes the transfer of 2,3-dihydroxybenzoate onto the phosphopantetheinylated (holo)EntB to yield the covalently arylated EntB. Finally, arylated EntB, ATP, and L-serine are used as substrates for the reaction catalyzed by EntF to generate enterobactin enterobactin + 6 AMP + 6 diphosphate
-
 ?
ATP + 2,3-dihydroxybenzoate + phosphopantetheinylated EntB Escherichia coli reaction of EntE arylated EntB + AMP + diphosphate
-
 ?
ATP + arylated EntB + L-serine Escherichia coli reaction of EntF enterobactin + AMP + diphosphate
-
 ?

Organism

Organism UniProt Comment Textmining
Escherichia coli P0ADI4 gene entB; gene entB
-
Escherichia coli P10378 gene entE; gene entE
-
Escherichia coli P11454 gene entF; gene entF
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged EntE from Escherichia coli strain BL21(DE3) by nickel affinity chromatography Escherichia coli

Reaction

Reaction Comment Organism Reaction ID
6 ATP + 3 2,3-dihydroxybenzoate + 3 L-serine = enterobactin + 6 AMP + 6 diphosphate bi-uni-uni-bi ping-pong kinetic mechanism Escherichia coli
a

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
6 ATP + 3 2,3-dihydroxybenzoate + 3 L-serine overall reaction. EntB, -D, -E, and -F are then required to catalyze the ATP-dependent assembly of enterobactin from three molecules each of 2,3-dihydroxybenzoate and L-serine. EntD, a phosphopantetheinyl transferase, uses coenzyme A to phosphopantetheinylate S245 of the aryl carrier protein domain of EntB. Next, EntE catalyzes the transfer of 2,3-dihydroxybenzoate onto the phosphopantetheinylated (holo) EntB to yield the covalently arylated EntB. Finally, arylated EntB, ATP, and L-serine are used as substrates for the reaction catalyzed by EntF to generate enterobactin Escherichia coli enterobactin + 6 AMP + 6 diphosphate
-
 ?
6 ATP + 3 2,3-dihydroxybenzoate + 3 L-serine overall reaction. EntB, -D, -E, and -F are then required to catalyze the ATP-dependent assembly of enterobactin from three molecules each of 2,3-dihydroxybenzoate and L-serine. EntD, a phosphopantetheinyl transferase, uses coenzyme A to phosphopantetheinylate S245 of the aryl carrier protein domain of EntB. Next, EntE catalyzes the transfer of 2,3-dihydroxybenzoate onto the phosphopantetheinylated (holo)EntB to yield the covalently arylated EntB. Finally, arylated EntB, ATP, and L-serine are used as substrates for the reaction catalyzed by EntF to generate enterobactin Escherichia coli enterobactin + 6 AMP + 6 diphosphate
-
 ?
ATP + 2,3-dihydroxybenzoate + phosphopantetheinylated EntB reaction of EntE Escherichia coli arylated EntB + AMP + diphosphate
-
 ?
ATP + 3-hydroxybenzoate + phosphopantetheinylated EntB reaction of EntE Escherichia coli arylated EntB + AMP + diphosphate
-
 ?
ATP + 4-aminosalicylic acid + phosphopantetheinylated EntB reaction of EntE Escherichia coli arylated EntB + AMP + diphosphate
-
 ?
ATP + arylated EntB + L-serine reaction of EntF Escherichia coli enterobactin + AMP + diphosphate
-
 ?
ATP + salicylic acid + phosphopantetheinylated EntB reaction of EntE Escherichia coli arylated EntB + AMP + diphosphate
-
 ?
additional information no activity with 4-aminobenzoic acid Escherichia coli ?
-
 ?

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
 assay at Escherichia coli

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.3
-
 3-hydroxybenzoate pH 7.8, 25°C, recombinant EntE Escherichia coli
0.8
-
 salicylic acid pH 7.8, 25°C, recombinant EntE Escherichia coli
2.8
-
 ATP pH 7.8, 25°C, recombinant EntE Escherichia coli
2.8
-
 2,3-Dihydroxybenzoate pH 7.8, 25°C, recombinant EntE Escherichia coli
2.8
-
 phosphopantetheinylated EntB pH 7.8, 25°C, recombinant EntE Escherichia coli
4.4
-
 4-aminosalicylic acid pH 7.8, 25°C, recombinant EntE Escherichia coli

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.8
-
 assay at Escherichia coli

Cofactor

Cofactor Comment Organism Structure
ATP
-
 Escherichia coli

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
additional information
-
 additional information thermodynamics and inhibition kinetics, stopped-flow measurements Escherichia coli

General Information

General Information Comment Organism
metabolism chorismate is converted to 2,3-dihydroxybenzoate via the sequential catalytic activities of EntC, -B, and -A Escherichia coli

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
4.6
-
 3-hydroxybenzoate pH 7.8, 25°C, recombinant EntE Escherichia coli
11
-
 salicylic acid pH 7.8, 25°C, recombinant EntE Escherichia coli
15
-
 4-aminosalicylic acid pH 7.8, 25°C, recombinant EntE Escherichia coli
390
-
 ATP pH 7.8, 25°C, recombinant EntE Escherichia coli
880
-
 2,3-Dihydroxybenzoate pH 7.8, 25°C, recombinant EntE Escherichia coli
980
-
 phosphopantetheinylated EntB pH 7.8, 25°C, recombinant EntE Escherichia coli