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Literature summary extracted from

  • Kataoka, K.; Takada, H.; Tanizawa, K.; Yoshimura, T.; Esaki, N.; Ohshima, T.; Soda, K.
    Construction and characterization of chimeric enzyme consisting of an amino-terminal domain of phenylalanine dehydrogenase and a carboxy-terminal domain of leucine dehydrogenase (1994), J. Biochem., 116, 931-936.
    View publication on PubMed

Application

EC Number Application Comment Organism
1.4.1.20 analysis the chimeric enzyme has a specific activity of 6% of that of the parental phenylalanine dehydrogenase Thermoactinomyces intermedius

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.4.1.9 chimeric enzyme consisting of an amino-terminal domain of phenylalanine dehydrogenase and a carboxy-terminal domain of leucine dehydrogenase Geobacillus stearothermophilus
1.4.1.20 chimeric enzyme consisting of the N-terminal domain of Thermoactinomyces intermedius phenylalanine dehydrogenase, containing the substrate-binding region and the C-terminal domain of leucine dehydrogenase from Bacillus stearothermophilus containing the NAD+-binding region, expression in Escherichia coli Thermoactinomyces intermedius

Protein Variants

EC Number Protein Variants Comment Organism
1.4.1.9 additional information chimeric enzyme consisting of an amino-terminal domain of phenylalanine dehydrogenase and a carboxy-terminal domain of leucine dehydrogenase containing the NAD+-binding region, the substrate specificity of the chimeric enzyme in the reductive amination is an admixture of those of the two parent enzymes Geobacillus stearothermophilus
1.4.1.20 additional information construction of a chimeric enzyme consisting of the N-terminal domain of Thermoactinomyces intermedius phenylalanine dehydrogenase, containing the substrate-binding region and the C-terminal domain of leucine dehydrogenase from Bacillus stearothermophilus containing the NAD+-binding region. The chimeric enzyme has a specific activity of 6% of that of the parental phenylalanine dehydrogenase and shows a broad substrate specificity in the oxidative deamination, like phenylalanine dehydrogenase. However, it acts much more effectively than phenylalanine dehydrogenase on Ile and Val Thermoactinomyces intermedius

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.4.1.20 0.06
-
 L-Leu chimeric enzyme Thermoactinomyces intermedius
1.4.1.20 0.11
-
 L-Phe
-
 Thermoactinomyces intermedius
1.4.1.20 0.12
-
 L-Phe chimeric enzyme Thermoactinomyces intermedius
1.4.1.20 0.17
-
 NAD+
-
 Thermoactinomyces intermedius

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
1.4.1.9 additional information
-
 MW of the chimeric enzyme consisting of an amino-terminal domain of phenylalanine dehydrogenase and a carboxy-terminal domain of leucine dehydrogenase is 72000 Da, determined by gel filtration Geobacillus stearothermophilus
1.4.1.9 40000
-
 2 * 40000, SDS-PAGE Geobacillus stearothermophilus
1.4.1.20 72000
-
 chimeric enzyme consisting of the N-terminal domain of Thermoactinomyces intermedius phenylalanine dehydrogenase, containing the substrate-binding region and the C-terminal domain of leucine dehydrogenase from Bacillus stearothermophilus containing the NAD+-binding region, gel filtration Thermoactinomyces intermedius

Organism

EC Number Organism UniProt Comment Textmining
1.4.1.9 Geobacillus stearothermophilus
-
-
-
1.4.1.20 Thermoactinomyces intermedius
-
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.4.1.9 chimeric enzyme consisting of an amino-terminal domain of phenylalanine dehydrogenase and a carboxy-terminal domain of leucine dehydrogenase Geobacillus stearothermophilus
1.4.1.20 construction of a chimeric enzyme consisting of the N-terminal domain of Thermoactinomyces intermedius phenylalanine dehydrogenase, containing the substrate-binding region and the C-terminal domain of leucine dehydrogenase from Bacillus stearothermophilus containing the NAD+-binding region Thermoactinomyces intermedius

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.4.1.9 L-Leu + H2O + NAD+
-
 Geobacillus stearothermophilus 4-methyl-2-oxopentanoate + NH3 + NADH
-
 r
1.4.1.9 additional information substrate specificity of the chimeric enzyme consisting of an amino-terminal domain of phenylalanine dehydrogenase and a carboxy-terminal domain of leucine dehydrogenase Geobacillus stearothermophilus ?
-
 ?
1.4.1.20 2-oxo-4-methylpentanoate + NH3 + NADH 16% of the activity with phenylpyruvate Thermoactinomyces intermedius L-Ile + NAD+ + H2O
-
 ?
1.4.1.20 2-oxo-4-methylthiobutanoate + NH3 + NADH 55% of the activity with phenylpyruvate Thermoactinomyces intermedius L-Met + NAD+ + H2O
-
 ?
1.4.1.20 2-oxobutanoate + NH3 + NADH 5.5% of the activity with phenylpyruvate Thermoactinomyces intermedius 2-aminobutanoate + NAD+ + H2O
-
 ?
1.4.1.20 2-oxohexanoate + NH3 + NADH 130% of the activity with phenylpyruvate Thermoactinomyces intermedius 2-aminohexanoate + H2O + NAD+
-
 ?
1.4.1.20 2-oxoisohexanoate + NH3 + NADH 47% of the activity with phenylpyruvate Thermoactinomyces intermedius L-Leu + NAD+ + H2O
-
 ?
1.4.1.20 2-oxopentanoate + NH3 + NADH 37% of the activity with phenylpyruvate Thermoactinomyces intermedius L-norvaline + NAD+ + H2O
-
 ?
1.4.1.20 4-hydroxyphenylpyruvate + NH3 + NADH 80% of the activity with phenylpyruvate Thermoactinomyces intermedius L-Tyr + NAD+ + H2O
-
 ?
1.4.1.20 alloisoleucine + H2O + NAD+ 26% of the activity with L-Phe Thermoactinomyces intermedius 3-methyl-2-oxopentanoate + NADH + NH3
-
 ?
1.4.1.20 L-Met + H2O + NAD+ 2.2% of the activity with L-Phe Thermoactinomyces intermedius 2-oxo-4-methylthiobutanoate + NH3 + NADH
-
 ?
1.4.1.20 L-norleucine + H2O + NAD+ 30% of the activity with L-Phe Thermoactinomyces intermedius 2-oxohexanoic acid + NADH + NH3
-
 ?
1.4.1.20 L-norvaline + H2O + NAD+ 28% of the activity with L-Phe Thermoactinomyces intermedius 2-oxopentanoate + NH3 + NADH
-
 ?
1.4.1.20 L-Phe + H2O + NAD+
-
 Thermoactinomyces intermedius phenylpyruvate + NH3 + NADH
-
 r
1.4.1.20 L-Tyr + H2O + NAD+ 40% of the activity with L-Phe Thermoactinomyces intermedius 4-hydroxyphenylpyruvate + NH3 + NADH
-
 ?
1.4.1.20 m-fluoro-DL-phenylalanine + H2O + NAD+ as effective as L-Phe Thermoactinomyces intermedius 3-(3-fluorophenyl)-2-oxopropionate + NADH + NH3
-
 ?
1.4.1.20 additional information specific activity of the chimeric enzyme is 6% of that of the parental phenylalanine dehydrogenase and shows a broad substrate specificity in the oxidative deamination, like phenylalanine dehydrogenase. However, it acts much more effectively than phenylalanine dehydrogenase on Ile and Val. The parent enzyme and the chimeric enzyme belong to the pro-S specific dehydrogenase Thermoactinomyces intermedius ?
-
 ?
1.4.1.20 o-fluoro-DL-phenylalanine + H2O + NAD+ 65% of the activity with L-Phe Thermoactinomyces intermedius 3-(2-fluorophenyl)-2-oxopropionate + NADH + NH3
-
 ?
1.4.1.20 p-amino-L-phenylalanine + H2O + NAD+
-
 Thermoactinomyces intermedius ?
-
 ?
1.4.1.20 p-fluoro-DL-phenylalanine + H2O + NAD+ 118% of the activity with L-Phe Thermoactinomyces intermedius (4-fluorophenyl)-2-oxopropionate + NADH + NH3
-
 ?

Subunits

EC Number Subunits Comment Organism
1.4.1.9 dimer 2 * 40000, SDS-PAGE Geobacillus stearothermophilus

Temperature Stability [°C]

EC Number Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
1.4.1.9 54
-
 pH 7.0-9.5, 60 min, chimeric enzyme consisting of an amino-terminal domain of phenylalanine dehydrogenase and a carboxy-terminal domain of leucine dehydrogenase, stable Geobacillus stearothermophilus
1.4.1.9 58
-
 pH 7.0-9.5, 60 min, chimeric enzyme consisting of an amino-terminal domain of phenylalanine dehydrogenase and a carboxy-terminal domain of leucine dehydrogenase, loss of activity Geobacillus stearothermophilus
1.4.1.20 54
-
 60 min, pH 7.0-9.5, chimeric enzyme is stable Thermoactinomyces intermedius
1.4.1.20 58
-
 pH 7.0-9.5, 60 min, chimeric enzyme, loss of activity above Thermoactinomyces intermedius
1.4.1.20 65 70 60 min, stable Thermoactinomyces intermedius

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.4.1.9 additional information
-
 pH-optima for the chimeric enzyme consisting of an amino-terminal domain of phenylalanine dehydrogenase and a carboxy-terminal domain of leucine dehydrogenase Geobacillus stearothermophilus
1.4.1.20 10.6
-
 reductive amination of phenylpyruvate, chimeric enzyme Thermoactinomyces intermedius
1.4.1.20 10.7 11 oxidative deamination of L-Phe, chimeric enzyme Thermoactinomyces intermedius

Cofactor

EC Number Cofactor Comment Organism Structure
1.4.1.9 NAD+
-
 Geobacillus stearothermophilus
1.4.1.9 NADH
-
 Geobacillus stearothermophilus
1.4.1.20 NAD+
-
 Thermoactinomyces intermedius
1.4.1.20 NADH
-
 Thermoactinomyces intermedius