BRENDA - Enzyme Database

Conversion of glutaconate CoA-transferase from Acidaminococcus fermentans into an acyl-CoA hydrolase by site-directed mutagenesis

Mack, M.; Buckel, W; FEBS Lett. 405, 209-212 (1997)

Data extracted from this reference:

Cloned(Commentary)
EC Number
Commentary
Organism
2.8.3.12
overexpression of betaE54N and betaE54A mutant enzyme in Escherichia coli, fusion of the genes gctA and gctB, encoding the 2 subunits of Gct, yields GctF, which is expressed in Escherichia coli as a 65 kDa protein with 30% of wild-type activity; overexpression of wild-type and betaE54D mutant enzyme in Escherichia coli
Acidaminococcus fermentans
3.1.2.1
expression in Escherichia coli
Acidaminococcus fermentans
Crystallization (Commentary)
EC Number
Crystallization
Organism
2.8.3.12
betaE54D mutant enzyme, expressed in Escherichia coli
Acidaminococcus fermentans
Engineering
EC Number
Amino acid exchange
Commentary
Organism
2.8.3.12
E54A
mutant without CoA-transferase or acyl-CoA hydrolase activity; mutation in the subunit GctB, inactive mutant
Acidaminococcus fermentans
2.8.3.12
E54D
mutation decreases CoA-transferase and increases acyl-CoA hydrolase activity, mechanism, mutant catalyses the hydrolysis of glutaryl-CoA, acetyl-CoA and 3-butenoyl-CoA, mutant Escherichia coli strain shows reduced growth rate after induction; mutation in the subunit GctB
Acidaminococcus fermentans
2.8.3.12
E54N
mutant without CoA-transferase or acyl-CoA hydrolase activity; mutation in the subunit GctB, inactive mutant
Acidaminococcus fermentans
KM Value [mM]
EC Number
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
2.8.3.12
0.028
-
glutaryl-CoA
pH 7, cosubstrate acetate, wild-type enzyme
Acidaminococcus fermentans
2.8.3.12
0.064
-
glutaryl-CoA
pH 7, cosubstrate acetate, 65 kDa GctF fusion protein
Acidaminococcus fermentans
Organism
EC Number
Organism
Primary Accession No. (UniProt)
Commentary
Textmining
2.8.3.12
Acidaminococcus fermentans
-
-
-
3.1.2.1
Acidaminococcus fermentans
-
-
-
Purification (Commentary)
EC Number
Commentary
Organism
2.8.3.12
betaE54D mutant enzyme and 65 kDa GctF fusion protein, expressed in Escherichia coli
Acidaminococcus fermentans
Reaction
EC Number
Reaction
Commentary
Organism
2.8.3.12
acetyl-CoA + (E)-glutaconate = acetate + glutaconyl-1-CoA
reaction mechanism
Acidaminococcus fermentans
Specific Activity [micromol/min/mg]
EC Number
Specific Activity Minimum [Ámol/min/mg]
Specific Activity Maximum [Ámol/min/mg]
Commentary
Organism
2.8.3.12
additional information
-
-
Acidaminococcus fermentans
Substrates and Products (Substrate)
EC Number
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
2.8.3.12
acetyl-CoA + (E)-glutaconate
-
94485
Acidaminococcus fermentans
acetate + glutaconyl-1-CoA
-
94485
Acidaminococcus fermentans
?
2.8.3.12
glutaryl-CoA + acetate
-
94485
Acidaminococcus fermentans
glutarate + acetyl-CoA
-
94485
Acidaminococcus fermentans
?
2.8.3.12
additional information
catalytic residue is E-54 of subunit B, catalytic mechanism
94485
Acidaminococcus fermentans
?
-
-
-
-
3.1.2.1
3-butenoyl-CoA
-
94485
Acidaminococcus fermentans
3-butenoate + CoA
-
-
-
?
3.1.2.1
glutaryl-CoA
-
94485
Acidaminococcus fermentans
glutarate + CoA
-
-
-
?
Subunits
EC Number
Subunits
Commentary
Organism
2.8.3.12
octamer
(alphabeta)4
Acidaminococcus fermentans
Turnover Number [1/s]
EC Number
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
2.8.3.12
additional information
-
additional information
-
Acidaminococcus fermentans
2.8.3.12
39
-
glutaryl-CoA
pH 7, cosubstrate acetate, 65 kDa GctF fusion protein
Acidaminococcus fermentans
2.8.3.12
140
-
glutaryl-CoA
pH 7, cosubstrate acetate, wild-type enzyme
Acidaminococcus fermentans
pH Optimum
EC Number
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
2.8.3.12
7
-
assay at
Acidaminococcus fermentans
Cloned(Commentary) (protein specific)
EC Number
Commentary
Organism
2.8.3.12
overexpression of betaE54N and betaE54A mutant enzyme in Escherichia coli, fusion of the genes gctA and gctB, encoding the 2 subunits of Gct, yields GctF, which is expressed in Escherichia coli as a 65 kDa protein with 30% of wild-type activity; overexpression of wild-type and betaE54D mutant enzyme in Escherichia coli
Acidaminococcus fermentans
3.1.2.1
expression in Escherichia coli
Acidaminococcus fermentans
Crystallization (Commentary) (protein specific)
EC Number
Crystallization
Organism
2.8.3.12
betaE54D mutant enzyme, expressed in Escherichia coli
Acidaminococcus fermentans
Engineering (protein specific)
EC Number
Amino acid exchange
Commentary
Organism
2.8.3.12
E54A
mutant without CoA-transferase or acyl-CoA hydrolase activity; mutation in the subunit GctB, inactive mutant
Acidaminococcus fermentans
2.8.3.12
E54D
mutation decreases CoA-transferase and increases acyl-CoA hydrolase activity, mechanism, mutant catalyses the hydrolysis of glutaryl-CoA, acetyl-CoA and 3-butenoyl-CoA, mutant Escherichia coli strain shows reduced growth rate after induction; mutation in the subunit GctB
Acidaminococcus fermentans
2.8.3.12
E54N
mutant without CoA-transferase or acyl-CoA hydrolase activity; mutation in the subunit GctB, inactive mutant
Acidaminococcus fermentans
KM Value [mM] (protein specific)
EC Number
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
2.8.3.12
0.028
-
glutaryl-CoA
pH 7, cosubstrate acetate, wild-type enzyme
Acidaminococcus fermentans
2.8.3.12
0.064
-
glutaryl-CoA
pH 7, cosubstrate acetate, 65 kDa GctF fusion protein
Acidaminococcus fermentans
Purification (Commentary) (protein specific)
EC Number
Commentary
Organism
2.8.3.12
betaE54D mutant enzyme and 65 kDa GctF fusion protein, expressed in Escherichia coli
Acidaminococcus fermentans
Specific Activity [micromol/min/mg] (protein specific)
EC Number
Specific Activity Minimum [Ámol/min/mg]
Specific Activity Maximum [Ámol/min/mg]
Commentary
Organism
2.8.3.12
additional information
-
-
Acidaminococcus fermentans
Substrates and Products (Substrate) (protein specific)
EC Number
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
2.8.3.12
acetyl-CoA + (E)-glutaconate
-
94485
Acidaminococcus fermentans
acetate + glutaconyl-1-CoA
-
94485
Acidaminococcus fermentans
?
2.8.3.12
glutaryl-CoA + acetate
-
94485
Acidaminococcus fermentans
glutarate + acetyl-CoA
-
94485
Acidaminococcus fermentans
?
2.8.3.12
additional information
catalytic residue is E-54 of subunit B, catalytic mechanism
94485
Acidaminococcus fermentans
?
-
-
-
-
3.1.2.1
3-butenoyl-CoA
-
94485
Acidaminococcus fermentans
3-butenoate + CoA
-
-
-
?
3.1.2.1
glutaryl-CoA
-
94485
Acidaminococcus fermentans
glutarate + CoA
-
-
-
?
Subunits (protein specific)
EC Number
Subunits
Commentary
Organism
2.8.3.12
octamer
(alphabeta)4
Acidaminococcus fermentans
Turnover Number [1/s] (protein specific)
EC Number
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
2.8.3.12
additional information
-
additional information
-
Acidaminococcus fermentans
2.8.3.12
39
-
glutaryl-CoA
pH 7, cosubstrate acetate, 65 kDa GctF fusion protein
Acidaminococcus fermentans
2.8.3.12
140
-
glutaryl-CoA
pH 7, cosubstrate acetate, wild-type enzyme
Acidaminococcus fermentans
pH Optimum (protein specific)
EC Number
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
2.8.3.12
7
-
assay at
Acidaminococcus fermentans