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Literature summary extracted from

  • Dawson, A.; Chen, M.; Fyfe, P.K.; Guo, Z.; Hunter, W.N.
    Structure and reactivity of Bacillus subtilis MenD catalyzing the first committed step in menaquinone biosynthesis (2010), J. Mol. Biol., 401, 253-264.
    View publication on PubMedView publication on EuropePMC

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
2.2.1.9 to 2.35 A resolution, in complex with thiamine diphosphate and Mn2+. basic residues Arg32, Arg106, Arg409, Arg428, and Lys299 interact with carboxylate and hydroxyl groups to align substrates for catalysis in combination with a cluster of the non-polar residues Ile489, Phe490, and Leu493 on one side of the active site. Arg409 plays a significant role in binding both substrates while Arg428 contributes mainly to binding of 2-oxoglutarate. Arg32 and in particular Arg106 are critical for recognition of isochorismate Bacillus subtilis

Protein Variants

EC Number Protein Variants Comment Organism
2.2.1.9 F490A 0.6% of wild-type activity Bacillus subtilis
2.2.1.9 I489A 1% of wild-type activity Bacillus subtilis
2.2.1.9 K299A 150% of wild-type activity Bacillus subtilis
2.2.1.9 L493A 80% of wild-type activity Bacillus subtilis
2.2.1.9 R106A mutation in the second subunit forming the active site, 10% of wild-type activity Bacillus subtilis
2.2.1.9 R32A mutation in the second subunit forming the active site, 30% of wild-type activity Bacillus subtilis
2.2.1.9 R409A 3% of wild-type activity Bacillus subtilis
2.2.1.9 R428A 1% of wild-type activity Bacillus subtilis

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
2.2.1.9 0.0077
-
 2-oxoglutarate mutant R106A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.018
-
 2-oxoglutarate mutant L493A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.022
-
 2-oxoglutarate mutant K299A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.022
-
 2-oxoglutarate mutant R32A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.022
-
 2-oxoglutarate wild-type, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.159
-
 2-oxoglutarate mutant R409A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.274
-
 2-oxoglutarate mutant F490A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.346
-
 2-oxoglutarate mutant I489A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.514
-
 2-oxoglutarate mutant R428A, pH 7.8, 23°C Bacillus subtilis

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
2.2.1.9 290000
-
 gel filtration Bacillus subtilis

Organism

EC Number Organism UniProt Comment Textmining
2.2.1.9 Bacillus subtilis P23970
-
-

Reaction

EC Number Reaction Comment Organism Reaction ID
2.2.1.9 isochorismate + 2-oxoglutarate = 5-enolpyruvoyl-6-hydroxy-2-succinyl-cyclohex-3-ene-1-carboxylate + CO2 two-stage mechanism that is primarily driven by the chemical properties of the cofactor thiamine diphosphate Bacillus subtilis
a

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.2.1.9 isochorismate + 2-oxoglutarate
-
 Bacillus subtilis 5-enolpyruvoyl-6-hydroxy-2-succinyl-cyclohex-3-ene-1-carboxylate + CO2
-
 ?

Subunits

EC Number Subunits Comment Organism
2.2.1.9 tetramer dimer of dimer, crystallization data, 4 x 64100, calculated Bacillus subtilis

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
2.2.1.9 0.01
-
 2-oxoglutarate mutant R106A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.02
-
 2-oxoglutarate mutant F490A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.05
-
 2-oxoglutarate mutant R409A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.06
-
 2-oxoglutarate mutant R428A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.08
-
 2-oxoglutarate mutant R32A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.1
-
 2-oxoglutarate mutant I489A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.17
-
 2-oxoglutarate mutant L493A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.23
-
 2-oxoglutarate wild-type, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.37
-
 2-oxoglutarate mutant K299A, pH 7.8, 23°C Bacillus subtilis

Cofactor

EC Number Cofactor Comment Organism Structure
2.2.1.9 thiamine diphosphate in absence of any residue that can act as a general acid/base the cofactor N4' atom is a critical component of the mechanism Bacillus subtilis

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
2.2.1.9 0.07
-
 2-oxoglutarate mutant F490A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.12
-
 2-oxoglutarate mutant R428A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 0.27
-
 2-oxoglutarate mutant I489A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 1.5
-
 2-oxoglutarate mutant R106A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 2.7
-
 2-oxoglutarate mutant R409A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 3.7
-
 2-oxoglutarate mutant R32A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 8.3
-
 2-oxoglutarate mutant L493A, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 10.7
-
 2-oxoglutarate wild-type, pH 7.8, 23°C Bacillus subtilis
2.2.1.9 16.7
-
 2-oxoglutarate mutant K299A, pH 7.8, 23°C Bacillus subtilis