Information on EC 1.4.1.20 - phenylalanine dehydrogenase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
1.4.1.20
-
RECOMMENDED NAME
GeneOntology No.
phenylalanine dehydrogenase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
L-phenylalanine + H2O + NAD+ = phenylpyruvate + NH3 + NADH + H+
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
oxidative deamination
-
-
-
-
redox reaction
-
-
-
-
reductive amination
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Tyrosine metabolism
-
-
Phenylalanine metabolism
-
-
Phenylalanine, tyrosine and tryptophan biosynthesis
-
-
Metabolic pathways
-
-
Biosynthesis of secondary metabolites
-
-
Biosynthesis of antibiotics
-
-
SYSTEMATIC NAME
IUBMB Comments
L-phenylalanine:NAD+ oxidoreductase (deaminating)
The enzymes from Bacillus badius and Sporosarcina ureae are highly specific for L-phenylalanine; that from Bacillus sphaericus also acts on L-tyrosine.
CAS REGISTRY NUMBER
COMMENTARY hide
69403-12-9
-
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
-
an inborn error of amino acid metabolism, in which the conversion of L-phenylalanine to L-tyrosine is impaired and can cause profound mental retardation if not detected and treated soon after birth, is phenylketonuria, a disease due to an autosomal recessive inheritance that codes for a type of phenylalanine hydroxylase with reduced enzymatic activity, resulting in high levels of L-Phe in inborn fluid and phenylpyruvate in urine
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
2-(3-hydroxy-1-adamantyl)-2-oxoethanoic acid + NH3 + NADH
(S)-3-hydroxyadamantylglycine + H2O + NAD+
show the reaction diagram
-
modified PDH which contains two amino acid changes at the C-terminus and a 12 amino acid extension of the C-terminus is more effective with keto acid 2-(3-hydroxy-1-adamantyl)-2-oxoethanoic acid and less effective with phenylpyruvate than the wild type PDH
-
-
?
2-chloro-phenylpyruvate + NH3 + NADH
2-chloro-L-phenylalanine + H2O + NAD+
show the reaction diagram
-
7% of activity with phenylpyruvate
-
-
?
2-fluoro-phenylpyruvate + NH3 + NADH
2-fluoro-L-phenylalanine + H2O + NAD+
show the reaction diagram
-
6.6% of activity with phenylpyruvate
-
-
?
2-oxo-4-methylpentanoate + NH3 + NADH
L-Ile + NAD+ + H2O
show the reaction diagram
2-oxo-4-methylthiobutanoate + NH3 + NADH
L-Met + NAD+ + H2O
show the reaction diagram
2-oxo-4-phenylbutanoic acid + NH3 + NADH + H+
(S)-2-amino-4-phenylbutyric acid + H2O + NAD+
show the reaction diagram
2-oxobutanoate + NH3 + NADH
2-aminobutanoate + NAD+ + H2O
show the reaction diagram
-
5.5% of the activity with phenylpyruvate
-
-
?
2-oxohexanoate + NH3 + NADH
2-aminohexanoate + H2O + NAD+
show the reaction diagram
2-oxoisohexanoate + NH3 + NADH
L-Leu + NAD+ + H2O
show the reaction diagram
2-oxopentanoate + NH3 + NADH
L-norvaline + NAD+ + H2O
show the reaction diagram
3-chloro-phenylpyruvate + NH3 + NADH
3-chloro-L-phenylalanine + H2O + NAD+
show the reaction diagram
-
0.8% of activity with phenylpyruvate
-
-
?
3-cyclohexyl-2-oxopropionic acid + NH3 + NADH
3-cyclohexyl-2-aminopropionic acid + H2O + NAD+
show the reaction diagram
-
6.1% of activity with phenylpyruvate
-
-
?
3-fluoro-phenylpyruvate + NH3 + NADH
3-fluoro-L-phenylalanine + H2O + NAD+
show the reaction diagram
-
23% of activity with phenylpyruvate
-
-
?
4-chloro-L-phenylalanine + H2O + NAD+
4-chloro-phenylpyruvate + NH3 + NADH
show the reaction diagram
-
-
-
-
?
4-chloro-phenylpyruvate + NH3 + NADH
4-chloro-L-phenylalanine + H2O + NAD+
show the reaction diagram
-
62% of activity with phenylpyruvate
-
-
?
4-fluoro-L-phenylalanine + H2O + NAD+
4-fluoro-phenylpyruvate + NH3 + NADH
show the reaction diagram
-
-
-
-
?
4-fluoro-phenylpyruvate + NH3 + NADH
4-fluoro-L-phenylalanine + H2O + NAD+
show the reaction diagram
4-hydroxyphenylpyruvate + NH3 + NADH
L-Tyr + NAD+ + H2O
show the reaction diagram
4-methoxy-L-phenylalanine + H2O + NAD+
4-methoxy-phenylpyruvate + NH3 + NADH
show the reaction diagram
-
-
-
-
?
4-methoxy-phenylpyruvate + NH3 + NADH
4-methoxy-L-phenylalanine + H2O + NAD+
show the reaction diagram
4-methyl-L-phenylalanine + H2O + NAD+
4-methyl-phenylpyruvate + NH3 + NADH
show the reaction diagram
-
-
-
-
?
4-methyl-phenylpyruvate + NH3 + NADH
4-methyl-L-phenylalanine + H2O + NAD+
show the reaction diagram
-
15% of activity with phenylpyruvate
-
-
?
4-nitro-L-phenylalanine + H2O + NAD+
4-nitro-phenylpyruvate + NH3 + NADH
show the reaction diagram
-
-
-
-
?
4-nitro-phenylpyruvate + NH3 + NADH
4-nitro-L-phenylalanine + H2O + NAD+
show the reaction diagram
-
reaction with the immobilized mutant enzyme N145A
-
-
?
4-pyridyl-phenylpyruvate + NH3 + NADH
4-pyridyl-L-phenylalanine + H2O + NAD+
show the reaction diagram
-
9% of activity with phenylpyruvate
-
-
?
4-trifluoromethyl-phenylpyruvate + NH3 + NADH
4-trifluoromethyl-L-phenylalanine + H2O + NAD+
show the reaction diagram
alloisoleucine + H2O + NAD+
3-methyl-2-oxopentanoate + NADH + NH3
show the reaction diagram
alpha-ketocaproic acid + NH3 + NADH
alpha-aminocaproic acid + H2O + NAD+
show the reaction diagram
-
12.8% of activity with phenylpyruvate
-
-
?
DL-propargylglycine + phenazine ethosulfate + 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyltetrazolium chloridehydrate + NAD+
? + NADH
show the reaction diagram
-
substrate is poorly fertilized by wild type enzyme, but using muteins with different mutations higher turnover can be achieved
-
-
?
indole-3-pyruvate + NH3 + NADH
L-Trp + NADH + H2O
show the reaction diagram
indolepyruvate + NH3 + NADH
?
show the reaction diagram
L-2-amino-n-butyric acid + H2O + NAD+
2-oxobutanoate + NADH + NH3
show the reaction diagram
-
1.0% of the activity with L-Phe
-
-
?
L-2-aminohexanoic acid + H2O + NAD+
2-oxohexanoate + NADH + NH3
show the reaction diagram
-
19% of the activity with L-Phe
-
-
?
L-alpha-amino-beta-phenylbutyrate + H2O + NAD+
2-oxo-3-phenylbutyrate + NADH + NH3
show the reaction diagram
-
7.0% of the activity with L-Phe
-
-
?
L-ethionine + H2O + NAD+
4-ethylthio-2-oxobutanoate + NH3 + NADH
show the reaction diagram
L-isoleucine + H2O + NAD+
3-methyl-2-oxopentanoate + NH3 + NADH
show the reaction diagram
-
-
-
-
r
L-Leu + H2O + NAD+
3-methyl-2-oxopentanoic acid + NH3 + NADH
show the reaction diagram
L-leucine + H2O + NAD+
3,3-dimethyl-2-oxo-butanoate + NH3 + NADH
show the reaction diagram
L-Met + H2O + NAD+
2-oxo-4-methylthiobutanoate + NH3 + NADH
show the reaction diagram
L-methionine + H2O + NAD+
(4-methylsulfanyl)-2-oxobutanoate + NH3 + NADH
show the reaction diagram
L-norleucine + H2O + NAD+
2-oxohexanoate + NH3 + NADH
show the reaction diagram
L-norleucine + H2O + NAD+
2-oxohexanoic acid + NADH + NH3
show the reaction diagram
L-norvaline + H2O + NAD+
2-oxopentanoate + NH3 + NADH
show the reaction diagram
L-Phe + H2O + NAD+
phenylpyruvate + NH3 + NADH
show the reaction diagram
L-phenylalanine + H2O + 3-acetylpyridine-NAD+
phenylpyruvate + NH3 + acetylpyridine-NADH
show the reaction diagram
L-phenylalanine + H2O + 3-pyridinealdehyde-NAD+
phenylpyruvate + NH3 + 3-pyridinealdehyde-NADH
show the reaction diagram
L-phenylalanine + H2O + deamino-NAD+
phenylpyruvate + NH3 + deamino-NADH
show the reaction diagram
L-phenylalanine + H2O + NAD+
phenylpyruvate + NH3 + NADH
show the reaction diagram
L-phenylalanine + H2O + NAD+
phenylpyruvate + NH3 + NADH + H+
show the reaction diagram
L-phenylalanine + H2O + NADP+
phenylpyruvate + NH3 + NADPH
show the reaction diagram
L-phenylalanine + H2O + oxidized beta-nicotinamide guanine dinucleotide
phenylpyruvate + NH3 + reduced beta-nicotinamide guanine dinucleotide
show the reaction diagram
-
at 86% of the activity with NAD+
-
?
L-phenylalanine + H2O + oxidized beta-nicotinamide hypoxanthine dinucleotide
phenylpyruvate + NH3 + reduced beta-nicotinamide hypoxanthine dinucleotide
show the reaction diagram
-
at 86% of the activity with NAD+
-
?
L-phenylalanine + H2O + thio-NAD+
phenylpyruvate + NH3 + thio-NADH
show the reaction diagram
-
at 86% of the activity with NAD+
-
?
L-phenylalanine + H2O + thionicotinamide-NAD+
phenylpyruvate + NH3 + thionicotinamide-NADH
show the reaction diagram
L-phenylalanine methyl ester + H2O + NAD+
phenylpyruvic acid methyl ester + NH3 + NADH
show the reaction diagram
L-phenylalaninol + H2O + NAD+
1-hydroxyacetone + NADH + NH3
show the reaction diagram
-
9.4% of the activity with L-Phe
-
-
?
L-Trp + H2O + NAD+
indole-3-pyruvate + NH3 + NADH
show the reaction diagram
L-tryptophan + H2O + NAD+
3-(1H-indol-3-yl)-2-oxopropanoate + NH3 + NADH
show the reaction diagram
-
2% of activity with L-phenylalanine
-
-
r
L-Tyr + H2O + NAD+
4-hydroxyphenylpyruvate + NH3 + NADH
show the reaction diagram
L-tyrosine + H2O + NAD+
(4-hydroxyphenyl)pyruvate + NH3 + NADH
show the reaction diagram
L-Val + H2O + NAD+
3-methyl-2-oxobutanoate + NH3 + NADH
show the reaction diagram
L-valine + H2O + NAD+
3-methyl-2-oxobutanoate + NH3 + NADH
show the reaction diagram
m-fluoro-DL-phenylalanine + H2O + NAD+
3-(3-fluorophenyl)-2-oxopropionate + NADH + NH3
show the reaction diagram
o-fluoro-DL-phenylalanine + H2O + NAD+
3-(2-fluorophenyl)-2-oxopropionate + NADH + NH3
show the reaction diagram
p-amino-L-phenylalanine + H2O + NAD+
?
show the reaction diagram
-
-
-
-
?
p-fluoro-DL-phenylalanine + H2O + NAD+
(4-fluorophenyl)-2-oxopropionate + NADH + NH3
show the reaction diagram
phenylalaninamide + H2O + NAD+
2-oxo-3-phenylpropionamide + NADH + NH3
show the reaction diagram
-
9.0% of the activity with L-Phe
-
-
?
phenylalanine hydroxamate + H2O + NAD+
2-oxo-3-phenylpropionic acid hydroxamate + NADH + NH3
show the reaction diagram
-
1.0% of the activity with L-Phe
-
-
?
phenylpyruvate + NH3 + NADH
L-Phe + H2O + NAD+
show the reaction diagram
-
-
-
-
r
phenylpyruvate + NH3 + NADH
L-phenylalanine + H2O + NAD+
show the reaction diagram
-
-
-
-
?
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
-
-
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
L-Phe + H2O + NAD+
phenylpyruvate + NH3 + NADH
show the reaction diagram
-
the enzyme is involved in the degradation of Phe
-
?
L-phenylalanine + H2O + NAD+
phenylpyruvate + NH3 + NADH
show the reaction diagram
L-phenylalanine + H2O + NAD+
phenylpyruvate + NH3 + NADH + H+
show the reaction diagram
phenylpyruvate + NH3 + NADH
L-phenylalanine + H2O + NAD+
show the reaction diagram
-
-
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
3-acetylpyridine-NAD+
-
241% of the activity with NAD+ in the reaction with L-Phe
3-pyridinealdehyde-NAD+
-
9.2% of the activity with NAD+ in the reaction with L-Phe
deamino-NAD+
-
86% of the activity with NAD+ in the reaction with L-Phe
thionicotinamide-NAD+
-
101% of the activity with NAD+ in the reaction with L-Phe
additional information
-
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
3-Phenylpropionate
-
competitive with L-Phe
4-hydroxyphenylethylamine
-
competitive with L-Phe
5,5'-dithio-bis(2-nitrobenzoic acid)
AgNO3
alpha-methyl-DL-phenylalanine
-
-
Cu2+
-
slight
D-Ethionine
-
-
D-Leu
D-Met
D-Norleucine
D-Phe
D-Trp
D-Tyr
D-Val
-
10 mM, 11% inhibition
HgCl2
L-Cys
-
10 mM, 41% inhibition
L-Ile
-
10 mM, 75% inhibition
L-Leu
L-Phe
L-phenylglycine
-
competitive with L-Phe
L-Phenyllactate
-
-
L-Val
-
10 mM, 23% inhibition
methyl acetyl phosphate
-
irreversible inactivation, simultaneous addition of substrate and coenzyme markedly protect from inactivation, the reagent can acetylate Lys69 and Lys81
NEM
-
1 mM: 79% inhibition
phenylenediamine
-
-
Phenylethylamine
phenylpropionate
-
-
phenylpyruvate
-
-
trans-cinnamate
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2.8
2-oxo-4-methyl-thiobutanoate
-
wild-type enzyme
13 - 50
2-oxo-4-methylpentanoate
2.1 - 87
2-oxo-4-methylthiobutanoate
2.44 - 47
2-Oxohexanoate
9.1 - 56
2-Oxoisohexanoate
35 - 107
2-Oxopentanoate
0.25 - 110
3-acetylpyridine-NAD+
50
3-pyridinealdehyde-NAD+
-
-
0.24 - 1.3
4-hydroxyphenylpyruvate
0.45
4-trifluoromethyl-phenylpyruvate
-
mutant enzyme N145A, at 25C inTris buffer 50mM pH 8.5, with NADH (0.1 mM), KCl (100 mM) and NH4Cl (400 mM)
11
alpha-keto-gamma-methiobutyrate
-
25C, pH 10.4
13
alpha-Ketocaproate
-
25C, pH 10.4
20
alpha-Ketoisocaproate
-
25C, pH 10.4
52
alpha-ketovalerate
-
25C, pH 10.4
0.18
deamino-NAD+
-
-
13
DL-alpha-keto-beta-methyl-n-valerate
-
25C, pH 10.4
2 - 298
DL-propargylglycine
7.7
Indolepyruvate
-
-
8
indolylpyruvate
-
-
0.27 - 4
L-ethionine
0.09 - 10
L-Ile
0.14
L-isoleucine
-
25C, pH 10.4
0.06 - 4.7
L-Leu
0.55
L-leucine
-
25C, pH 10.4
0.34 - 7.5
L-Met
2.7
L-methionine
-
25C, pH 10.4
0.79 - 2.1
L-norleucine
0.53 - 10
L-norvaline
0.09 - 140
L-Phe
0.018 - 9.6
L-phenylalanine
11
L-Trp
-
-
1.1 - 3.1
L-Tyr
0.48 - 8.6
L-tyrosine
2
L-valine
-
25C, pH 10.4
0.057 - 90
NAD+
0.017 - 0.13
NADH
78 - 96
NH3
387
NH4+
-
-
0.22
norleucine
-
wild-type enzyme
170
oxidized beta-nicotinamide guanine dinucleotide
-
-
40
oxidized beta-nicotinamide hypoxanthine dinucleotide
-
-
0.19
p-hydroxy-phenylpyruvate
-
25C, pH 10.4
0.34 - 2.4
p-hydroxyphenylpyruvate
0.052 - 23
phenylpyruvate
60
thioNAD+
-
-
0.18
thionicotinamide-NAD+
-
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
26
2-oxo-4-methylpentanoate
Thermoactinomyces intermedius
-
wild-type enzyme
37
2-oxo-4-methylthiobutanoate
Thermoactinomyces intermedius
-
wild-type enzyme
79
2-Oxohexanoate
Thermoactinomyces intermedius
-
wild-type enzyme
57
2-Oxoisohexanoate
Thermoactinomyces intermedius
-
wild-type enzyme
82
2-Oxopentanoate
Thermoactinomyces intermedius
-
wild-type enzyme
27
alpha-keto-gamma-methiobutyrate
Lysinibacillus sphaericus
-
25C, pH 10.4
51
alpha-Ketocaproate
Lysinibacillus sphaericus
-
25C, pH 10.4
27
alpha-Ketoisocaproate
Lysinibacillus sphaericus
-
25C, pH 10.4
33
alpha-ketovalerate
Lysinibacillus sphaericus
-
25C, pH 10.4
7.2
DL-alpha-keto-beta-methyl-n-valerate
Lysinibacillus sphaericus
-
25C, pH 10.4
0.5 - 1.82
DL-propargylglycine
0.1 - 1.1
L-ethionine
0.24 - 0.28
L-Ile
0.14
L-isoleucine
Lysinibacillus sphaericus
-
25C, pH 10.4
0.21 - 3.6
L-Leu
0.7
L-leucine
Lysinibacillus sphaericus
-
25C, pH 10.4
0.16 - 0.84
L-Met
1.2
L-methionine
Lysinibacillus sphaericus
-
25C, pH 10.4
0.17 - 3.2
L-norleucine
0.15 - 1.4
L-norvaline
0.75 - 816
L-Phe
0.608 - 68.01
L-phenylalanine
34 - 68
L-tyrosine
0.91
L-valine
Lysinibacillus sphaericus
-
25C, pH 10.4
90
p-hydroxy-phenylpyruvate
Lysinibacillus sphaericus
-
25C, pH 10.4
91 - 113
phenylpyruvate
additional information
additional information
Thermoactinomyces intermedius
-
turnover numbers of wild-type and mutant enzymes
-
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0054 - 0.04
DL-propargylglycine
937
0.33 - 279
L-phenylalanine
104
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2.6
3-Phenylpropionate
-
-
13.7
4-hydroxyphenylethylamine
-
-
20.8
alpha-methyl-DL-phenylalanine
-
-
1.76
D-Ethionine
-
-
0.14
D-Phe
-
-
1.58
D-Tyr
-
-
8.32
L-phenylglycine
-
-
1.27
NAD+
-
-
0.002
NADH
-
-
17.85
Phe
-
-
4.36
phenylenediamine
-
-
1.52
Phenylethylamine
-
-
0.07 - 7.34
phenylpyruvate
12.7
trans-cinnamic acid
-
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
9.1
-
native enzyme, pH 10.4, 30C
285.4
-
recombinant PheDH, oxidative deamination
450
-
PheDH after glycosidation
553.6
-
after gel filtration protocol
816
-
unmodified PheDH
4231
-
after polymer/salt aqueous two-phase systems (ATPS) protocol
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.5 - 9.5
-
reductive amination, immobilized enzyme
8.4 - 9
-
modified PDH which contains two amino acid changes at the C-terminus and a 12 amino acid extension of the C-terminus
8.5
-
reductive amination of phenylpyruvate, crude extract
9.3
-
reductive amination
9.7 - 10.1
-
mutant enzyme F124M/V125S/H126I/A127I/A128Y/R129Q, oxidative deamination
9.8
-
reductive amination of phenylpyruvate
10.2
-
reductive amination
10.3
-
reductive amination of phenylpyruvate
10.6
-
reductive amination of phenylpyruvate, chimeric enzyme
10.7 - 11
-
oxidative deamination of L-Phe, chimeric enzyme
10.8
-
oxidative deamination of L-Phe
11 - 12
-
native enzyme
11.3
-
oxidative deamination of L-Phe
12
-
oxidative deamination and reductive amination
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
9.3 - 10.2
-
pH 9.3: about 40% of maximal activity, pH 10.2: about 35% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
45 - 55
-
immobilized enzyme
55
-
neoglycoenzyme
60
-
immobilized enzyme
additional information
-
the enzyme glycosylated with mono-6-amino-6-deoxy-beta-cyclodextrin and mono-6-(5-carboxypentane-1-carboxamidoyl)-6-deoxy-beta-cyclodextrin contains about 18 mol and 15 mol oligosaccharide per mol of protein and retains 60% and 81% of the initial activity, respectively. The optimum temperature for the catalytic activity is increased in 10C after attaching the cyclodextrin residues
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20 - 70
-
20C: about 50% of maximal activity, 70C: about 70% of maximal activity
25 - 55
-
20C: about 60% of maximal activity, 55C, about 65% of maximal activity, native enzyme
25 - 60
-
20C: about 60% of maximal activity, 55C: about 85% of maximal activity, 60C: about 35% of maximal activity
30 - 70
-
-
40 - 53
-
40C: about 65% of maximal activity, 53C: optimum, no activity at 55C
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
40000
-
subunit of modified PDH (contains two amino acid changes at the C-terminus and a 12 amino acid extension of the C-terminus), SDS-PAGE
42000
-
gel filtration
55000
-
mutant enzyme F124M/V125S/H126I/A127I/A128Y/R129Q, monomeric enzyme form, gel filtration
70000
-
gel filtration
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
110000
-
mutant enzyme F124M/V125S/H126I/A127I/A128Y/R129Q, dimeric enzyme form, gel filtration
150000
-
gel filtration
290000
-
gel filtration
305000
-
equilibrium sedimentation
310000 - 340000
-
gel filtration
310000
-
gel filtration
325600
-
native enzyme, gel filtration
326000
-
SDS-PAGE, unmodified PheDH
330000
-
gel filtration
340000
342000
-
dextran-modified enzyme, gel filtration
360000
-
gel filtration
380000
-
gel filtration
720000
-
SDS-PAGE, neoglyoenzyme
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
decamer
hexamer
-
6 * 41000, wild-type enzyme, SDS-PAGE
monomer
octamer
tetramer
-
4 * 39500, SDS-PAGE
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycoprotein
-
the neoglycoenzyme retains 63% of its initial activity and contains about 2.5 mol of O-carboxymethyl poly-beta-cyclodextrin per mole of enzyme
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
hanging-drop method of vapour diffusion using ammonium sulfate as the precipitant. Two crystal forms are obtained in the presence and absence of the enzyme substrates phenylpyruvate or Phe and its coenzyme NADH
-
enzyme-NAD+-phenylpyruvate complex and enzyme-NAD+-beta-phenylpropionate complex
-
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4
-
30C, 1 h, less than 20% loss of activity
349634
7.5
-
native enzyme, 15 min, 96% activity remaining at 30-45C
723965
8 - 10.4
-
in pH 8.0 buffer with either 10% methanol or 10% ethanol L307V PheDH retains almost 85% original activity after 7 days, therafter activity continues to decrease gradually, most rapidly in pH 10.4 buffer with 10% ethanol, wild type PheDH is similarly stable, mutant N145A PheDH shows 68% retention of activity after 7 days at pH 8.0 with 10% methanol
686436
8 - 12.5
-
30C, 30 min, stable
349650
10.4
-
native enzyme, 15 min, no activity remaining at 30-45C
723965
11.3
-
30C, 1 h, stable
349634
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 45
-
native enzyme, 15 min, 96% activity remaining at pH 7.5, no activity remaining at pH 10.4
30
-
pH 9.0, stable
37
-
t1/2: 48 min, in absence of substrate or coenzyme. t1/2: 2000 min, in presence of 10 mM L-Leu and 1.0 mM NAD+
45 - 61
-
the optimum temperature for the glycosylated enzyme is increased by 15C and its thermostability is improved by about 6C over 10 min incubation, the conjugate is more resistant to thermal inactivation at different temperatures, ranging from 45 to 60C, the T50 value is increased from 54 to 61C for PheDH after glycosidation
53
-
pH 6.0, 5 min, 50% loss of activity; pH 9.5-10, 2 h, 50% loss of activity; phenylpyruvate destabilizes the activity, 1 M sorbitol or 1 M glycerol stabilize
58
-
pH 7.0-9.5, 60 min, chimeric enzyme, loss of activity above
62
-
T50-value of the dextran-modified enzyme
65 - 70
70
-
60 min, wild-type enzyme is stable
additional information
-
the enzyme glycosylated with mono-6-amino-6-deoxy-beta-cyclodextrin and mono-6-(5-carboxypentane-1-carboxamidoyl)-6-deoxy-beta-cyclodextrin contains about 18 mol and 15 mol oligosaccharide per mol of protein and retains 60% and 81% of the initial activity, respectively. the thermostability profile of the enzyme is improved abd its resistance to thermal inactivation at different temperatures ranging from 45C to 60C is noticeably increased after glycosylation
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
2.5 mM L-Phe after 21 days of incubation at 4C in 50 mM sodium phosphate buffer, pH 7.0
-
20% glycerol, 0.25 M sodium malonate or 0.5 M sodium glutarate stabilize the enzyme during storage at 4C
-
glycosylation increases temperature stability
-
PheDH covalently immobilized on amino-activated cellulose membrane by cross-linking with glutaraldehyde retains 82% of the initial activity, the enzyme-coated electrode retains full electrocatalytic activity after 16 days of storage at 4C in 50 mM sodium phosphate buffer (pH 7.0)
-
the enzyme chemically glycosylated with an end-group aminated dextran shows improved thermal and conformational stability
-
the enzyme-coated Au-CD electrode is highly stable, retaining about 97% of its initial electrocatalytic response toward
-
unstable in absence of glycerol and salt
-
ORGANIC SOLVENT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ethanol
-
Vmax values decrease substantially in the presence of 10% ethanol
Methanol
-
Vmax values decrease substantially in the presence of 10% (v/v) methanol
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
4C, 0.01 M potassium phosphate buffer, pH 7.0, 0.1 mM EDTA, 5 mM 2-mercaptoethanol, stable for at least 2 years in crystalline form
4C, 0.25 M sodium malonate, stable for 1 month
-
4C, immobilized on amino-activated cellulose membrane in 50 mM sodium phosphate buffer (pH 7.0), 16 days, no loss of activity
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
best purification results of recombinant PheDH are achieved in an polymer/salt aqueous two-phase systems (ATPS) with 9% of phenazine ethosulfate with a molecular mass of 6000 Da (PEG-6000) and 16% of K2HPO4
-
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
-
in aqueous two-phase systems composed of polyethylene glycol 6000 and ammonium sulfate with 8.5% (w/w) PEG-6000, 17.5% (w/w) (NH4)2SO4 and 13% (w/w) NaCl at pH 8.0
-
mutant enzyme F124M/V125S/H126I/A127I/A128Y/R129Q
-
Ni-affinity and anion exchange column chromatography
-
Ni-affinity column chromatography
-
Ni-affinity column chromatography and anion exchange column chromatography
-
partial
-
partial purification by Superdex 200 gel filtration
-
polyethylene glycol and ammonium sulfate aequous two-phase purification system with 9% (w/w) PEG-6000 and 16% (w/w) ammonium sulfate at pH 8.0
-
recombinant His-tagged enzyme from Escherchia coli by nickel affinity chromatography
-
recombinant PheDH, ammonium sulfate, DEAE-Toyopearl, hydroxyapatite, Sephadex G-200
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
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
-
espressed as recombinant protein in Escherichia coli strain BL21 (DE3)
-
expressed at high levels in Escherichia coli
-
expressed in Escherichia coli BL21 (DE3) cells
-
expressed in Escherichia coli BL21 (DE3)/pBBNH cells
-
expressed in Escherichia coli BL21(DE3) cells
-
expressed in Escherichia coli BL21(DE3)/pBBNH cells
-
expressed in Escherichia coli BL21/pBBNH cells
-
expressed in Escherichia coli via plasmids
-
expression in Bacillus subtilis
-
expression in Escherichia coli
-
gene pdh, DNA and amino acid sequence determination and analysis, overexpression of His-tagged enzyme in Escherchia coli, method optimization, overview
-
overexpression in Escherichia coli
-
the modified enzyme (contains two amino acid changes at the C-terminus and a 12 amino acid extension of the C-terminus) is expressed in Pichia pastoris strain SMD1168 or Escherichia coli strain SC 16496
-
wild-type and mutant enzymes L307V, G124A and G124A/L307V expressed in Escherichia coli
-
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D126N
-
mutant without enzymatic activity
D209G
-
mutant named 1stA7, active with DL-propargylglycine
D209G/Q18H/I336F
-
mutant named 1stA7/1stB6, active with DL-propargylglycine
E313G
-
mutant named H7H10, active with DL-propargylglycine
F110Y/G124C/G293A
-
mutant named H14A12, active with DL-propargylglycine
G124A/E313G
-
mutant named 25B12, active with DL-propargylglycine
G124A/E313N
-
mutant named 20E1, active with DL-propargylglycine
G124A/L307V
K224Q/L283F
-
mutant named 2ndB10, active with DL-propargylglycine
K224Q/L283F/E313G
-
mutant named 2ndB10/H7H10, active with DL-propargylglycine
N145I
-
19% of wild-type kcat with phenylalanine
Q18H/I336F
-
mutant named 1stB6, active with DL-propargylglycine
Q18H/I336F/E313G
-
mutant named 1stB6/H7H10, active with DL-propargylglycine
V135I/I308N/Q363H
-
mutant named H21D1, active with DL-propargylglycine
V33A/A206D/L283F
-
mutant named 2ndC2., active with DL-propargylglycine
F124M/V125S/H126I/A127I/A128Y/R129Q
-
the catalytic efficiencies of the mutant enzyme with aliphatic amino acids and aliphatic keto acids as substrates are 0.5% to 2% of that of the wild-type enzyme. The efficiencies for L-Phe and phenylpyruvate decreases to 0.0008% and 0.035% of that of the wild-type enzyme, respectively. Enzyme exists as monomeric or dimeric form, compared to wild-type enzyme which exists as hexameric enzyme form. Thermostability is lowered by mutation
K173A
-
37C, t1/2 of the mutant enzyme is 60 min, compared to 48 min for the wild type enzyme, without addition of substrate or cofactor
K69A
-
37C, t1/2 of the mutant enzyme is 50 min, compared to 48 min for the wild type enzyme, without addition of substrate or cofactor. Km-value for L-Phe is 1400fold higher compared to wild type enzyme, Km-value for phenylpyruvate is 128fold higher compared to wild type enzyme. Turnover number for deamination is 686fold lower than that of wild-type enzyme, turnover-number for amination is 43fold lower than that of wild-type enzyme
K69A/K81A
-
37C, t1/2 of the mutant enzyme is 450 min, compared to 48 min for the wild type enzyme, without addition of substrate or cofactor. Km-value for L-Phe is 200fold higher compared to wild type enzyme, Km-value for phenylpyruvate is 108fold higher compared to wild type enzyme. Turnover number for deamination is 110fold lower than that of wild-type enzyme, turnover-number for amination is 61fold lower than that of wild-type enzyme
K81A
-
37C, t1/2 of the mutant enzyme is 38 min, compared to 48 min for the wild type enzyme, without addition of substrate or cofactor. Turnover number for deamination is 440fold lower than that of wild-type enzyme, turnover-number for amination is 42fold lower than that of wild-type enzyme
K89A
-
37C, t1/2 of the mutant enzyme is 75 min, compared to 48 min for the wild type enzyme, without addition of substrate or cofactor
K90A
-
37C, t1/2 of the mutant enzyme is 80 min, compared to 48 min for the wild type enzyme, without addition of substrate or cofactor
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
biotechnology
diagnostics
-
the immobilized enzyme is useful for establishing an analytical spectrophotometric determination method of L-Phe in the diagnostics of phenyketonuria, an inborn error of amino acid metabolism in which the conversion of L-phenylalanine to L-tyrosine is impaired and can cause profound mental retardation if not detected and treated soon after birth. Early quantitative measurement of the plasma L-Phe is essential for the diagnosis of phenylketonuria and the control of dietary therapy of the patients. The conversion efficiency of the reactor with immobillized enzyme is 100% in the range of 0.005-0.6 mM Phe at 9 mM NAD+ with a total flow rate of 0.1 mL/min. The reactor is used for the analyses of 30 samples each for 3 h per day. The half-life period of the reactor is 15 days, method evaluation, overview
medicine
-
plays an important role in detection and screening of phenylketonuria diseases
synthesis