BRENDA - Enzyme Database

Residues Asn214, Gln211, Glu219 and Gln221 contained in the subfamily 3 catalytic signature of the isocitrate lyase from Pseudomonas aeruginosa are involved in its catalytic and thermal properties

Campos-Garcia, J.; Diaz-Perez, C.; Diaz-Perez, A.L.; World J. Microbiol. Biotechnol. 29, 991-999 (2013)

Data extracted from this reference:

Cloned(Commentary)
EC Number
Commentary
Organism
4.1.3.1
gene aceA, phylogenetic analysis, expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
Pseudomonas aeruginosa
Engineering
EC Number
Amino acid exchange
Commentary
Organism
4.1.3.1
E219A
site-directed mutagenesis of the highly conserved residue in the ICL subfamily 3 catalytic pattern, the mutant enzyme shows reduced activity compared to the wild-type enzyme
Pseudomonas aeruginosa
4.1.3.1
N214D
site-directed mutagenesis of the highly conserved residue in the ICL subfamily 3 catalytic pattern, the mutant enzyme shows 10fold reduced activity compared to the wild-type enzyme
Pseudomonas aeruginosa
4.1.3.1
Q211H
site-directed mutagenesis of the highly conserved residue in the ICL subfamily 3 catalytic pattern, the mutant enzyme shows reduced activity and elevated temperature optimum compared to the wild-type enzyme
Pseudomonas aeruginosa
4.1.3.1
Q221K
site-directed mutagenesis of the highly conserved residue in the ICL subfamily 3 catalytic pattern, the mutant enzyme shows reduced activity and elevated temperature optimum compared to the wild-type enzyme
Pseudomonas aeruginosa
KM Value [mM]
EC Number
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
4.1.3.1
additional information
-
additional information
Michaelis-Menten kinetics of wild-type and mutant enzyme's overview
Pseudomonas aeruginosa
4.1.3.1
0.25
-
isocitrate
recombinant His-tagged mutant N214D, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
0.29
-
isocitrate
recombinant His-tagged mutant Q211H, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
0.44
-
isocitrate
recombinant His-tagged wild-type enzyme, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
0.47
-
isocitrate
recombinant His-tagged mutant Q221K, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
0.94
-
isocitrate
recombinant His-tagged mutant E219A, pH 7.8, 30°C
Pseudomonas aeruginosa
Natural Substrates/ Products (Substrates)
EC Number
Natural Substrates
Organism
Commentary (Nat. Sub.)
Natural Products
Commentary (Nat. Pro.)
Organism (Nat. Pro.)
Reversibility
4.1.3.1
isocitrate
Pseudomonas aeruginosa
-
succinate + glyoxylate
-
-
?
Organism
EC Number
Organism
Primary Accession No. (UniProt)
Commentary
Textmining
4.1.3.1
Pseudomonas aeruginosa
Q9I0K4
gene aceA
-
Purification (Commentary)
EC Number
Commentary
Organism
4.1.3.1
ecombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography
Pseudomonas aeruginosa
Substrates and Products (Substrate)
EC Number
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
4.1.3.1
isocitrate
-
728845
Pseudomonas aeruginosa
succinate + glyoxylate
-
-
-
?
Temperature Optimum [°C]
EC Number
Temperature Optimum [°C]
Temperature Optimum Maximum [°C]
Commentary
Organism
4.1.3.1
30
-
wild-type enzyme and mutant E219A
Pseudomonas aeruginosa
4.1.3.1
36
-
mutant Q221K
Pseudomonas aeruginosa
4.1.3.1
39
-
mutant Q211H
Pseudomonas aeruginosa
Temperature Stability [°C]
EC Number
Temperature Stability Minimum [°C]
Temperature Stability Maximum [°C]
Commentary
Organism
4.1.3.1
45
-
purified enzyme, half-life of the wild-type enzyme is 40 min, half-lives of mutants E219A, Q211H, and Q221K are 25 min, 12 min, and 15 min
Pseudomonas aeruginosa
Turnover Number [1/s]
EC Number
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
4.1.3.1
0.56
-
isocitrate
recombinant His-tagged mutant N214D, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
3.25
-
isocitrate
recombinant His-tagged mutant Q221K, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
3.46
-
isocitrate
recombinant His-tagged mutant Q211H, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
4.63
-
isocitrate
recombinant His-tagged mutant E219A, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
9.81
-
isocitrate
recombinant His-tagged wild-type enzyme, pH 7.8, 30°C
Pseudomonas aeruginosa
pH Optimum
EC Number
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
4.1.3.1
7.6
-
-
Pseudomonas aeruginosa
Cloned(Commentary) (protein specific)
EC Number
Commentary
Organism
4.1.3.1
gene aceA, phylogenetic analysis, expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
Pseudomonas aeruginosa
Engineering (protein specific)
EC Number
Amino acid exchange
Commentary
Organism
4.1.3.1
E219A
site-directed mutagenesis of the highly conserved residue in the ICL subfamily 3 catalytic pattern, the mutant enzyme shows reduced activity compared to the wild-type enzyme
Pseudomonas aeruginosa
4.1.3.1
N214D
site-directed mutagenesis of the highly conserved residue in the ICL subfamily 3 catalytic pattern, the mutant enzyme shows 10fold reduced activity compared to the wild-type enzyme
Pseudomonas aeruginosa
4.1.3.1
Q211H
site-directed mutagenesis of the highly conserved residue in the ICL subfamily 3 catalytic pattern, the mutant enzyme shows reduced activity and elevated temperature optimum compared to the wild-type enzyme
Pseudomonas aeruginosa
4.1.3.1
Q221K
site-directed mutagenesis of the highly conserved residue in the ICL subfamily 3 catalytic pattern, the mutant enzyme shows reduced activity and elevated temperature optimum compared to the wild-type enzyme
Pseudomonas aeruginosa
KM Value [mM] (protein specific)
EC Number
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
4.1.3.1
additional information
-
additional information
Michaelis-Menten kinetics of wild-type and mutant enzyme's overview
Pseudomonas aeruginosa
4.1.3.1
0.25
-
isocitrate
recombinant His-tagged mutant N214D, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
0.29
-
isocitrate
recombinant His-tagged mutant Q211H, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
0.44
-
isocitrate
recombinant His-tagged wild-type enzyme, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
0.47
-
isocitrate
recombinant His-tagged mutant Q221K, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
0.94
-
isocitrate
recombinant His-tagged mutant E219A, pH 7.8, 30°C
Pseudomonas aeruginosa
Natural Substrates/ Products (Substrates) (protein specific)
EC Number
Natural Substrates
Organism
Commentary (Nat. Sub.)
Natural Products
Commentary (Nat. Pro.)
Organism (Nat. Pro.)
Reversibility
4.1.3.1
isocitrate
Pseudomonas aeruginosa
-
succinate + glyoxylate
-
-
?
Purification (Commentary) (protein specific)
EC Number
Commentary
Organism
4.1.3.1
ecombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography
Pseudomonas aeruginosa
Substrates and Products (Substrate) (protein specific)
EC Number
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
4.1.3.1
isocitrate
-
728845
Pseudomonas aeruginosa
succinate + glyoxylate
-
-
-
?
Temperature Optimum [°C] (protein specific)
EC Number
Temperature Optimum [°C]
Temperature Optimum Maximum [°C]
Commentary
Organism
4.1.3.1
30
-
wild-type enzyme and mutant E219A
Pseudomonas aeruginosa
4.1.3.1
36
-
mutant Q221K
Pseudomonas aeruginosa
4.1.3.1
39
-
mutant Q211H
Pseudomonas aeruginosa
Temperature Stability [°C] (protein specific)
EC Number
Temperature Stability Minimum [°C]
Temperature Stability Maximum [°C]
Commentary
Organism
4.1.3.1
45
-
purified enzyme, half-life of the wild-type enzyme is 40 min, half-lives of mutants E219A, Q211H, and Q221K are 25 min, 12 min, and 15 min
Pseudomonas aeruginosa
Turnover Number [1/s] (protein specific)
EC Number
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
4.1.3.1
0.56
-
isocitrate
recombinant His-tagged mutant N214D, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
3.25
-
isocitrate
recombinant His-tagged mutant Q221K, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
3.46
-
isocitrate
recombinant His-tagged mutant Q211H, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
4.63
-
isocitrate
recombinant His-tagged mutant E219A, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
9.81
-
isocitrate
recombinant His-tagged wild-type enzyme, pH 7.8, 30°C
Pseudomonas aeruginosa
pH Optimum (protein specific)
EC Number
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
4.1.3.1
7.6
-
-
Pseudomonas aeruginosa
General Information
EC Number
General Information
Commentary
Organism
4.1.3.1
evolution
the ICL family includes five subfamilies, and the 2-methylisocitrate lyase (MICL) family. The ICL from Pseudomonas aeruginosa (ICL-Pa) is identified in a different ICL node (subfamily 3) than other Pseudomonas ICL enzymes, phylogenetic analysis
Pseudomonas aeruginosa
4.1.3.1
additional information
conserved residues in the subfamily 3 signature of ICL-Pa play important roles in catalysis and thermostability and are likely associated with the catalytic loop structural conformation. Three-dimensional structural homology modeling of ICL-Pa wild-type and mutants, structure comparisons, active site modeling, overview. Residue N214 plays an essential role in ICL-Pa catalytic activity
Pseudomonas aeruginosa
4.1.3.1
physiological function
isocitrate lyase plays an important role in the ability of Pseudomonas aeruginosa to grow on fatty acids, acetate, acyclic terpenes, and amino acids
Pseudomonas aeruginosa
General Information (protein specific)
EC Number
General Information
Commentary
Organism
4.1.3.1
evolution
the ICL family includes five subfamilies, and the 2-methylisocitrate lyase (MICL) family. The ICL from Pseudomonas aeruginosa (ICL-Pa) is identified in a different ICL node (subfamily 3) than other Pseudomonas ICL enzymes, phylogenetic analysis
Pseudomonas aeruginosa
4.1.3.1
additional information
conserved residues in the subfamily 3 signature of ICL-Pa play important roles in catalysis and thermostability and are likely associated with the catalytic loop structural conformation. Three-dimensional structural homology modeling of ICL-Pa wild-type and mutants, structure comparisons, active site modeling, overview. Residue N214 plays an essential role in ICL-Pa catalytic activity
Pseudomonas aeruginosa
4.1.3.1
physiological function
isocitrate lyase plays an important role in the ability of Pseudomonas aeruginosa to grow on fatty acids, acetate, acyclic terpenes, and amino acids
Pseudomonas aeruginosa
KCat/KM [mM/s]
EC Number
kcat/KM Value [1/mMs-1]
kcat/KM Value Maximum [1/mMs-1]
Substrate
Commentary
Organism
Structure
4.1.3.1
2.8
-
isocitrate
recombinant His-tagged mutant N214D, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
4.9
-
isocitrate
recombinant His-tagged mutant E219A, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
6.9
-
isocitrate
recombinant His-tagged mutant Q221K, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
11.9
-
isocitrate
recombinant His-tagged mutant Q211H, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
22.3
-
isocitrate
recombinant His-tagged wild-type enzyme, pH 7.8, 30°C
Pseudomonas aeruginosa
KCat/KM [mM/s] (protein specific)
EC Number
KCat/KM Value [1/mMs-1]
KCat/KM Value Maximum [1/mMs-1]
Substrate
Commentary
Organism
Structure
4.1.3.1
2.8
-
isocitrate
recombinant His-tagged mutant N214D, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
4.9
-
isocitrate
recombinant His-tagged mutant E219A, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
6.9
-
isocitrate
recombinant His-tagged mutant Q221K, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
11.9
-
isocitrate
recombinant His-tagged mutant Q211H, pH 7.8, 30°C
Pseudomonas aeruginosa
4.1.3.1
22.3
-
isocitrate
recombinant His-tagged wild-type enzyme, pH 7.8, 30°C
Pseudomonas aeruginosa