BRENDA - Enzyme Database

Adenosylcobalamin-dependent glutamate mutase: examination of substrate and coenzyme binding in an engineered fusion protein possessing simplified subunit structure and kinetic properties

Chen, H.P.; Marsh, E.N.G.; Biochemistry 36, 14939-14945 (1997)

Data extracted from this reference:

Engineering
EC Number
Amino acid exchange
Commentary
Organism
5.4.99.1
additional information
the S subunit is genetically fused to the C-terminus of the E subunit through an 11 amino acid 5-Gly linker segment. The affinity of adenosylcobalamine is unchanged, but the turnover-number and the Km-value for Glu in the conversion of L-Glu to (2S,3S)-3-methylaspartate are decreased by about a third
Clostridium cochlearium
KM Value [mM]
EC Number
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
5.4.99.1
0.14
-
3-methylaspartate
genetically engineered enzyme with S subunit fused to the C-terminus of the E-subunit through an 11 amino acid (Gly-Gln)5-Gly linker segment
Clostridium cochlearium
5.4.99.1
0.58
-
L-Glu
genetically engineered enzyme with S subunit fused to the C-terminus of the E-subunit through an 11 amino acid (Gly-Gln)5-Gly linker segment
Clostridium cochlearium
Organism
EC Number
Organism
Primary Accession No. (UniProt)
Commentary
Textmining
5.4.99.1
Clostridium cochlearium
-
recombinant enzyme expressed in Escherichia coli
-
Substrates and Products (Substrate)
EC Number
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
5.4.99.1
L-Glu
-
3455
Clostridium cochlearium
threo-3-Methylaspartate
(2S,3S)-3-methylaspartate
3455
Clostridium cochlearium
-
Turnover Number [1/s]
EC Number
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
5.4.99.1
5.8
-
(2S,3S)-3-methylaspartate
genetically engineered enzyme with S subunit fused to the C-terminus of the E subunit through an 11 amino acid (Gly-Gln)5-Gly linker segment
Clostridium cochlearium
5.4.99.1
5.8
-
L-Glu
genetically engineered enzyme with S subunit fused to the C-terminus of the E subunit through an 11 amino acid (Gly-Gln)5-Gly linker segment
Clostridium cochlearium
pH Optimum
EC Number
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
5.4.99.1
7
-
protein with the S subunit genetically fused to the C-terminus of the E-subunit through an 11 amino acid (Gly-Gln)5-Gly linker segment
Clostridium cochlearium
5.4.99.1
8
8.5
wild type enzyme
Clostridium cochlearium
Cofactor
EC Number
Cofactor
Commentary
Organism
Structure
5.4.99.1
Cobalamin
the weakly associated subunits E and S combine to form the coenzyme binding site. Interactions between the protein and the adenosyl moiety do not serve to weaken the cobalt-carbon bond in the ground state, variation of the apparent Km-value for adenosylcobalamine with protein concentration. Km: 0.0055 mM in the reaction with L-Glu, Km: 0.0031 mM in the reaction with (2S,3S)-3-methylaspartate, genetically engineered enzyme with S subunit fused to the C-terminus of the E subunit through an 11 amino acid (Gly-Gln)5-Gly linker segment
Clostridium cochlearium
Cofactor (protein specific)
EC Number
Cofactor
Commentary
Organism
Structure
5.4.99.1
Cobalamin
the weakly associated subunits E and S combine to form the coenzyme binding site. Interactions between the protein and the adenosyl moiety do not serve to weaken the cobalt-carbon bond in the ground state, variation of the apparent Km-value for adenosylcobalamine with protein concentration. Km: 0.0055 mM in the reaction with L-Glu, Km: 0.0031 mM in the reaction with (2S,3S)-3-methylaspartate, genetically engineered enzyme with S subunit fused to the C-terminus of the E subunit through an 11 amino acid (Gly-Gln)5-Gly linker segment
Clostridium cochlearium
Engineering (protein specific)
EC Number
Amino acid exchange
Commentary
Organism
5.4.99.1
additional information
the S subunit is genetically fused to the C-terminus of the E subunit through an 11 amino acid 5-Gly linker segment. The affinity of adenosylcobalamine is unchanged, but the turnover-number and the Km-value for Glu in the conversion of L-Glu to (2S,3S)-3-methylaspartate are decreased by about a third
Clostridium cochlearium
KM Value [mM] (protein specific)
EC Number
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
5.4.99.1
0.14
-
3-methylaspartate
genetically engineered enzyme with S subunit fused to the C-terminus of the E-subunit through an 11 amino acid (Gly-Gln)5-Gly linker segment
Clostridium cochlearium
5.4.99.1
0.58
-
L-Glu
genetically engineered enzyme with S subunit fused to the C-terminus of the E-subunit through an 11 amino acid (Gly-Gln)5-Gly linker segment
Clostridium cochlearium
Substrates and Products (Substrate) (protein specific)
EC Number
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
5.4.99.1
L-Glu
-
3455
Clostridium cochlearium
threo-3-Methylaspartate
(2S,3S)-3-methylaspartate
3455
Clostridium cochlearium
-
Turnover Number [1/s] (protein specific)
EC Number
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
5.4.99.1
5.8
-
(2S,3S)-3-methylaspartate
genetically engineered enzyme with S subunit fused to the C-terminus of the E subunit through an 11 amino acid (Gly-Gln)5-Gly linker segment
Clostridium cochlearium
5.4.99.1
5.8
-
L-Glu
genetically engineered enzyme with S subunit fused to the C-terminus of the E subunit through an 11 amino acid (Gly-Gln)5-Gly linker segment
Clostridium cochlearium
pH Optimum (protein specific)
EC Number
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
5.4.99.1
7
-
protein with the S subunit genetically fused to the C-terminus of the E-subunit through an 11 amino acid (Gly-Gln)5-Gly linker segment
Clostridium cochlearium
5.4.99.1
8
8.5
wild type enzyme
Clostridium cochlearium