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Information on EC 1.4.3.3 - D-amino-acid oxidase and Organism(s) Trigonopsis variabilis and UniProt Accession Q99042
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1.4.3.3 D-amino-acid oxidaseIUBMB Comments
A flavoprotein (FAD). Wide specificity for D-amino acids. Also acts on glycine.
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Word Map
- 1.4.3.3
- d-serine
-
schizophrenia
- peroxisomal
- flavin
- n-methyl-d-aspartate
- d-alanine
- catalase
- fad
- nmda
-
deamination
- benzoate
-
flavoenzyme
- flavoproteins
- racemase
- variabilis
-
l-amino
-
neurotransmission
- d-aspartate
- gracilis
-
co-agonist
- rhodotorula
- cephalosporin
-
glutamatergic
- urate
- d-proline
- d-ala
- d-ser
-
imino
-
antipsychotic
-
hypofunction
- d-methionine
- isoalloxazine
- acylase
-
fad-containing
- toruloides
- rhodosporidium
- d-glutamate
-
fad-dependent
- d-cysteine
-
kynurenic
- sulfurtransferase
- d-valine
- synthesis
- medicine
- d-leucine
- cerium
-
7-aminocephalosporanic
- d-tryptophan
- d-phenylalanine
- neuregulin
- 3-mercaptopyruvate
- sarcosine
- industry
- biotechnology
- analysis
- diagnostics
- drug development
- pharmacology
The taxonomic range for the selected organisms is: Trigonopsis variabilis
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota, Archaea
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota, Archaea
Synonyms
d-amino acid oxidase, d-amino-acid oxidase, hdaao, d-aao, rgdaao, tvdao, tvdaao, d-aminoacid oxidase, peg-dao, pkdaao, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
D-amino acid oxidase
-
-
D-aminoacid oxidase
-
-
-
-
DAAO
DAMOX
-
-
-
-
DAO
ophio-amino-acid oxidase
-
-
-
-
oxidase, D-amino acid
-
-
-
-
DAAO
-
-
-
-
DAAO
-
-
DAO
-
-
-
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
redox reaction
-
-
-
-
oxidation
-
-
-
-
reduction
-
-
-
-
oxidative deamination
oxidative deamination
-
-
-
-
PATHWAY SOURCE
PATHWAYS
BRENDA
KEGG
-
-, -
SYSTEMATIC NAME
IUBMB Comments
D-amino-acid:oxygen oxidoreductase (deaminating)
A flavoprotein (FAD). Wide specificity for D-amino acids. Also acts on glycine.
CAS REGISTRY NUMBER
COMMENTARY
9000-88-8
-
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
D-alanine + H2O + O2
pyruvate + NH3 + H2O2
-
-
-
?
D-serine + H2O + O2
2-oxo-3-hydroxypropionate + NH3 + H2O2
-
-
-
?
D-tryptophan + H2O + O2
indol-3-pyruvic acid + NH3 + H2O2
-
-
-
?
cephalosporin C
alpha-ketoadipinyl-7-aminocephalosporanic acid
-
-
-
-
?
cephalosporin C + H2O + O2
7-aminocephalosporanic acid + ? + H2O2
conversion of cephalosporin C to 7-aminocephalosporanic acid and spontaneous decarboxylation of oxoadipyl-7-amino cephalosporanic acid is promoted by the H2O2 formed in the oxidase reaction of TvDAO
-
-
?
D-Arg + H2O + O2
5-guanidino-2-oxopentanoate + NH3 + H2O2
-
43% of the activity with D-Val
-
-
?
D-arginine + H2O + O2
5-guanidino-2-oxopentanoate + NH3 + H2O2
-
-
-
-
r
D-asparagine + H2O + O2
2-oxosuccinamic acid + NH3 + H2O2
-
67% activity compared to D-alanine
-
-
?
D-aspartate + H2O + O2
oxaloacetate + NH3 + H2O2
-
2% activity compared to D-alanine
-
-
?
D-citrulline + H2O + O2
2-oxo-5-ureidopentanoate + NH3 + H2O2
-
-
-
-
r
D-citrulline + H2O + O2
2-oxo-5-ureidopentanoic acid + NH3 + H2O2
-
-
-
-
?
D-cysteine + H2O + O2
2-oxo-3-thiopropionic acid + NH3 + H2O2
-
9% activity compared to D-alanine
-
-
?
D-ethionine + H2O + O2
4-ethylsulfanyl-2-oxobutyric acid + NH3 + H2O2
-
-
-
-
?
D-Gln + H2O + O2
2-oxoglutaramate + NH3 + H2O2
-
81% of the activity with D-Val
-
-
?
D-Glu + H2O + O2
alpha-ketoglutarate + NH3 + H2O2
-
9% of the activity with D-Val
-
-
?
D-glutamate + H2O + O2
alpha-ketoglutarate + NH3 + H2O2
-
9% activity compared to D-alanine
-
-
?
D-glutamine + H2O + O2
2-oxoglutaramate + NH3 + H2O2
-
78% activity compared to D-alanine
-
-
?
D-His + H2O + O2
3-(1H-imidazol-4-yl)-2-oxopropanoic acid + NH3 + H2O2
-
88% of the activity with D-Val
-
-
?
D-Ile + H2O + O2
3-methyl-2-oxopentanoate + NH3 + H2O2
-
76% of the activity with D-Val
-
-
?
D-isoleucine + H2O + O2
3-methyl-2-oxopentanoate + NH3 + H2O2
-
69% activity compared to D-alanine
-
-
?
D-isoleucine + H2O + O2
3-methyl-2-oxopentanoic acid + NH3 + H2O2
-
-
-
-
?
D-Leu + H2O + O2
4-methyl-2-oxopentanoate + NH3 + H2O2
-
32% of the activity with D-Val
-
-
?
D-Met + H2O + O2
4-methylthio-2-oxobutanoic acid + NH3 + H2O2
-
best substrate
-
-
?
D-methionine + 2,6-dichloroindophenol
4-methylsulfanyl-2-oxobutanoate + reduced 2,6-dichloroindophenol
-
-
-
-
?
D-methionine + H2O + O2
4-methylthio-2-oxobutanoate + NH3 + H2O2
-
highly active
-
-
?
D-Ser + H2O + O2
2-oxo-3-hydroxypropionate + NH3 + H2O2
-
22% of the activity with D-Val
-
-
?
D-serine + H2O + O2
2-oxo-3-hydroxypropionate + NH3 + H2O2
-
poor substrate
-
-
?
D-threonine + H2O + O2
2-oxo-3-hydroxybutyric acid + NH3 + H2O2
-
2% activity compared to D-alanine
-
-
?
D-tryptophan + H2O + O2
indol-3-pyruvate + NH3 + H2O2
-
129% activity compared to D-alanine
-
-
?
D-tryptophan + H2O + O2
indol-3-pyruvic acid + NH3 + H2O2
-
-
-
-
?
D-Val + H2O + O2
alpha-ketoisovalerate + NH3 + H2O2
-
best substrate
-
-
?
D-valine + H2O + O2
alpha-ketoisovalerate + NH3 + H2O2
-
highly active
-
-
?
cephalosporin C + H2O + O2
7-(5-oxoadipoamido)cephalosporanic acid + NH3 + H2O2
-
-
-
?
cephalosporin C + H2O + O2
7-(5-oxoadipoamido)cephalosporanic acid + NH3 + H2O2
-
-
-
-
?
cephalosporin C + H2O + O2
7-(5-oxoadipoamido)cephalosporanic acid + NH3 + H2O2
-
-
-
?
D-methionine + H2O + O2
4-methylthio-2-oxobutanoic acid + NH3 + H2O2
best substrate
-
-
?
D-methionine + H2O + O2
4-methylthio-2-oxobutanoic acid + NH3 + H2O2
by far best substrate
-
-
?
cephalosporin C + H2O + O2
7-(5-oxoadipoamido)cephalosporanic acid + NH3 + H2O2
-
-
-
-
?
cephalosporin C + H2O + O2
7-(5-oxoadipoamido)cephalosporanic acid + NH3 + H2O2
-
13% of the activity with D-Val
-
-
?
cephalosporin C + H2O + O2
7-(5-oxoadipoamido)cephalosporanic acid + NH3 + H2O2
-
14% activity compared to D-alanine
-
-
?
D-Ala + H2O + O2
pyruvate + NH3 + H2O2
-
97% of the activity with D-Val
-
-
?
D-amino acid + H2O + O2
2-oxo acid + NH3 + H2O2
via formation of an imino acid
-
-
?
D-amino acid + H2O + O2
2-oxocarboxylate + NH3 + H2O2
-
-
-
-
?
D-arginine + H2O + O2
5-guanidino-2-oxopentanoic acid + NH3 + H2O2
-
-
-
-
?
D-arginine + H2O + O2
5-guanidino-2-oxopentanoic acid + NH3 + H2O2
-
42% activity compared to D-alanine
-
-
?
D-Asn + H2O + O2
2-oxosuccinamate + NH3 + H2O2
-
65% of the activity with D-Val
-
-
?
D-histidine + H2O + O2
3-(1H-imidazol-4-yl)-2-oxopropanoate + NH3 + H2O2
-
-
-
-
?
D-histidine + H2O + O2
3-(1H-imidazol-4-yl)-2-oxopropanoate + NH3 + H2O2
-
69% activity compared to D-alanine
-
-
?
D-leucine + H2O + O2
4-methyl-2-oxopentanoic acid + NH3 + H2O2
-
-
-
-
?
D-leucine + H2O + O2
4-methyl-2-oxopentanoic acid + NH3 + H2O2
-
37% activity compared to D-alanine
-
-
?
D-lysine + H2O + O2
6-amino-2-oxohexanoic acid + NH3 + H2O2
-
poor substrate
-
-
?
D-lysine + H2O + O2
6-amino-2-oxohexanoic acid + NH3 + H2O2
-
7% activity compared to D-alanine
-
-
?
D-Met + H2O + O2
4-methylthio-2-oxobutanoate + NH3 + H2O2
-
78% of the ativity with D-Val
-
-
?
D-methionine + H2O + O2
4-methylsulfanyl-2-oxobutanoate + NH3 + H2O2
-
-
-
-
?
D-methionine + H2O + O2
4-methylsulfanyl-2-oxobutanoate + NH3 + H2O2
-
with 6-dichloroindophenol as electron acceptor
-
-
?
D-methionine + H2O + O2
4-methylthio-2-oxobutanoic acid + NH3 + H2O2
-
-
-
-
?
D-methionine + H2O + O2
4-methylthio-2-oxobutanoic acid + NH3 + H2O2
-
77% activity compared to D-alanine
-
-
?
D-Phe + H2O + O2
phenylpyruvate + NH3 + H2O2
-
36% of the activity with D-Val
-
-
?
D-phenylalanine + H2O + O2
phenylpyruvate + NH3 + H2O2
-
44% activity compared to D-alanine
-
-
?
D-phenylalanine + H2O + O2
phenylpyruvic acid + NH3 + H2O2
-
-
-
-
?
D-phenylalanine + H2O + O2
phenylpyruvic acid + NH3 + H2O2
-
activity of the DAAO after incubation in water-insoluble ionic liquids is higher than in water-soluble ones
-
-
?
D-proline + H2O + O2
2-oxopentanoic acid + NH3 + H2O2
-
25% of the activity with D-Val
-
-
?
D-proline + H2O + O2
2-oxopentanoic acid + NH3 + H2O2
-
21% activity compared to D-alanine
-
-
?
D-serine + H2O + O2
2-oxo-3-hydroxypropionic acid + NH3 + H2O2
-
-
-
-
?
D-serine + H2O + O2
2-oxo-3-hydroxypropionic acid + NH3 + H2O2
-
23% activity compared to D-alanine
-
-
?
D-threonine + H2O + O2
2-oxo-3-hydroxybutyrate + NH3 + H2O2
-
poor substrate
-
-
?
D-threonine + H2O + O2
2-oxo-3-hydroxybutyrate + NH3 + H2O2
-
4% of the activity with D-Val
-
-
?
D-Trp + H2O + O2
indol-3-pyruvate + NH3 + H2O2
-
38% of the activity with D-val
-
-
?
D-Tyr + H2O + O2
4-hydroxyphenylpyruvic acid + NH3 + H2O2
-
17% of the activity with D-Val
-
-
?
D-tyrosine + H2O + O2
4-hydroxyphenylpyruvic acid + NH3 + H2O2
-
-
-
-
?
D-tyrosine + H2O + O2
4-hydroxyphenylpyruvic acid + NH3 + H2O2
-
19% activity compared to D-alanine
-
-
?
D-valine + H2O + O2
alpha-ketoisovaleric acid + NH3 + H2O2
-
-
-
-
?
D-valine + H2O + O2
alpha-ketoisovaleric acid + NH3 + H2O2
-
63% activity compared to D-alanine
-
-
?
glycine + 2 H2O + O2
2 formic acid + NH3 + H2O2
-
poor substrate
-
-
?
glycine + 2 H2O + O2
2 formic acid + NH3 + H2O2
-
2% activity compared to D-alanine
-
-
?
additional information
?
-
best substrate is by far D-methionine followed by D-phenylalanine, D-tryptophan, D-valin, and D-alanine
-
-
?
additional information
?
-
D-aspartate and D-glutamate are no substrates
-
-
?
additional information
?
-
DAAOs can be divided into two groups regarding their substrate specificity, the first group prefers amino acids with small apolar side chains (D-Ala is the best substrate), the second group prefers D-amino acids possessing large hydrophobic side chains such as D-Trp, D-Met, D-Val, and D-Phe, usually the small amino acid Gly and the charged (acidic or basic) amino acids are poor DAAO substrates
-
-
?
additional information
?
-
the mechanism of the enzyme regulation is complex and multi-parametric because the same enzyme simultaneously influences the level of different D-amino acids, which can result in opposing effects, overview
-
-
?
additional information
?
-
the enzyme catalyzes oxidative deamination of D-amino acids yielding hydrogen peroxide and an imino acid. The latter is further non-enzymatically hydrolyzed to an alpha-keto acid and ammonium. DAAO is highly specific towards D-isomers of amino acids, it is almost inactive towards the corresponding L-isomer. The wild-type enzyme is inactive towards D-Asp, being however very active with D-Ala
-
-
?
additional information
?
-
-
nearly inactive toward D-aspartic and D-glutamic acids
-
-
?
additional information
?
-
-
the enzyme is involved in the conversion of cephalosporin C
-
-
?
additional information
?
-
conversion of cephalosporine C to 7-aminocephalosporanic acid and spontaneous decarboxylation of oxoadipyl-7-amino cephalosporanic acid is promoted by the H2O2 formed in the oxidase reaction of TvDAO
-
-
?
additional information
?
-
-
conversion of cephalosporine C to 7-aminocephalosporanic acid and spontaneous decarboxylation of oxoadipyl-7-amino cephalosporanic acid is promoted by the H2O2 formed in the oxidase reaction of TvDAO
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
D-serine + H2O + O2
2-oxo-3-hydroxypropionate + NH3 + H2O2
-
-
-
?
cephalosporin C + H2O + O2
7-(5-oxoadipoamido)cephalosporanic acid + NH3 + H2O2
-
-
-
-
?
D-arginine + H2O + O2
5-guanidino-2-oxopentanoic acid + NH3 + H2O2
-
-
-
-
?
D-citrulline + H2O + O2
2-oxo-5-ureidopentanoic acid + NH3 + H2O2
-
-
-
-
?
D-ethionine + H2O + O2
4-ethylsulfanyl-2-oxobutyric acid + NH3 + H2O2
-
-
-
-
?
D-histidine + H2O + O2
3-(1H-imidazol-4-yl)-2-oxopropanoate + NH3 + H2O2
-
-
-
-
?
D-isoleucine + H2O + O2
3-methyl-2-oxopentanoic acid + NH3 + H2O2
-
-
-
-
?
D-leucine + H2O + O2
4-methyl-2-oxopentanoic acid + NH3 + H2O2
-
-
-
-
?
D-methionine + H2O + O2
4-methylsulfanyl-2-oxobutanoate + NH3 + H2O2
-
-
-
-
?
D-methionine + H2O + O2
4-methylthio-2-oxobutanoic acid + NH3 + H2O2
-
-
-
-
?
D-phenylalanine + H2O + O2
phenylpyruvic acid + NH3 + H2O2
-
-
-
-
?
D-serine + H2O + O2
2-oxo-3-hydroxypropionic acid + NH3 + H2O2
-
-
-
-
?
D-tryptophan + H2O + O2
indol-3-pyruvic acid + NH3 + H2O2
-
-
-
-
?
D-tyrosine + H2O + O2
4-hydroxyphenylpyruvic acid + NH3 + H2O2
-
-
-
-
?
D-valine + H2O + O2
alpha-ketoisovaleric acid + NH3 + H2O2
-
-
-
-
?
D-amino acid + H2O + O2
2-oxocarboxylate + NH3 + H2O2
-
-
-
-
?
additional information
?
-
the mechanism of the enzyme regulation is complex and multi-parametric because the same enzyme simultaneously influences the level of different D-amino acids, which can result in opposing effects, overview
-
-
?
additional information
?
-
-
the enzyme is involved in the conversion of cephalosporin C
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
FAD
contains one molecule of noncovalently bound FAD per subunit
FAD
-
-
391857, 391858, 654786, 671491, 672709, 685721, 685750, 685758, 696959, 711306, 724401, 724887, 726433, 726525
FAD
-
added FAD does not stabilize DAO at and below 35°C, but it contributes up to 3.6fold extra stability to the enzyme activity at temperatures higher than 35°C
FAD
-
dependent, dissociation of FAD is a main contributor to the loss of activity
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
poly(methylene-co-guanidine)
-
in the presence of poly(methylene-co-guanidine) (1.8%, w/v) the enzyme activity decreases irreversibly with a half-life time of about 1.75 min at 25°C
-
additional information
-
oxidative modification of Trigonopsis variabilis D-amino acid oxidase in vivo is traceable as the conversion of Cys108 into a stable cysteine sulfinic acid, causes substantial loss of activity and thermostability of the enzyme
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
-
activity of the DAAO after incubation in water-insoluble ionic liquids is higher than in water-soluble ones
-
additional information
-
exogenous FAD (0.01 to 1 mM) added to assay or protein storage buffers does not enhance the catalytic activity of native and recombinant DAO
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
29.3
D-Lys
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
36.6
D-Ser
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
11.1
D-Thr
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
5.5
D-alanine
apparent value, reconstituted holoenzyme at pH 8.0 and 30°C
6
cephalosporin C
-
immobilized enzyme, in 100 mM potassium phosphate buffer, pH 8.0, at 25°C
9
cephalosporin C
-
free enzyme, in 100 mM potassium phosphate buffer, pH 8.0, at 25°C
16.7
D-Ala
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
3
D-alanine
-
free enzyme, in 100 mM potassium phosphate buffer, pH 8.0, at 25°C
4
D-alanine
-
immobilized enzyme, in 100 mM potassium phosphate buffer, pH 8.0, at 25°C
22
D-Asn
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
22.6
D-Asn
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
29
D-Asn
-
mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
50
D-Asn
-
Km above 50 mM, mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.227
D-Leu
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.78
D-Leu
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
1.19
D-Leu
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
1.81
D-Leu
-
mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.46
D-Met
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
2.9
D-Met
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
3.3
D-Met
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
4.2
D-Met
-
mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.068
D-Phe
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.134
D-Phe
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.37
D-Phe
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
1.68
D-Phe
-
mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.45
D-Trp
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.46
D-Trp
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.49
D-Trp
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
2.7
D-Trp
-
mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.087
D-Tyr
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.45
D-Tyr
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.84
D-Tyr
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
14.4
D-Val
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
41
D-Val
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
46
D-Val
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
3.54
D-Lys
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
20.5
D-Ser
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
1.75
D-Thr
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
95
D-alanine
apparent value, reconstituted holoenzyme at pH 8.0 and 30°C
3 - 6
cephalosporin C
-
immobilized enzyme, in 100 mM potassium phosphate buffer, pH 8.0, at 25°C
6.167
cephalosporin C
recombinant wild-type enzyme, pH 7.5, 22°C
55
cephalosporin C
-
free enzyme, in 100 mM potassium phosphate buffer, pH 8.0, at 25°C
108.6
D-Ala
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
44
D-alanine
-
immobilized enzyme, in 100 mM potassium phosphate buffer, pH 8.0, at 25°C
74
D-alanine
-
free enzyme, in 100 mM potassium phosphate buffer, pH 8.0, at 25°C
0.7
D-Asn
-
Km less than 0.7 s-1, mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
13
D-Asn
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
50
D-Asn
-
mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
62.4
D-Asn
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
29.1
D-Leu
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
46
D-Leu
-
mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
56
D-Leu
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
66.6
D-Leu
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
42
D-Met
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
63
D-Met
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
80.5
D-Met
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
89
D-Met
-
mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.84
D-methionine
-
recombinant mutant C108S, pH 7.5, 37°C, with 6-dichloroindophenolate
1.5
D-methionine
-
recombinant wild-type enzyme, pH 7.5, 37°C, with 6-dichloroindophenolate
2.8
D-methionine
-
recombinant mutant C108D, pH 7.5, 37°C, with 6-dichloroindophenolate
200
D-methionine
-
recombinant wild-type enzyme, pH 7.5, 37°C, with O2
27.2
D-Phe
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
47.1
D-Phe
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
58.8
D-Phe
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
63
D-Phe
-
mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
7.7
D-Trp
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
31
D-Trp
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
33
D-Trp
-
mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
42.4
D-Trp
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
15.9
D-Tyr
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
22.5
D-Tyr
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
72
D-Tyr
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
11.5
D-Val
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
14.6
D-Val
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
85.3
D-Val
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
6.5
D-Ala
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.12
D-Lys
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.56
D-Ser
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.16
D-Thr
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
3.83
cephalosporin C
recombinant wild-type enzyme, pH 7.5, 22°C
0.59
D-Asn
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
1.7
D-Asn
-
mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
2.8
D-Asn
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
14.8
D-Asn
-
Km less than 14.8 mM-1 s-1, mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
25
D-Leu
-
mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
37.3
D-Leu
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
56
D-Leu
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
245
D-Leu
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
12.7
D-Met
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
21
D-Met
-
mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
22
D-Met
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
175
D-Met
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
37
D-Phe
-
mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
73.9
D-Phe
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
351
D-Phe
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
865
D-Phe
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
12
D-Trp
-
mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
17
D-Trp
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
67.3
D-Trp
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
86.5
D-Trp
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
50
D-Tyr
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
85.8
D-Tyr
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
183
D-Tyr
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.25
D-Val
-
mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
0.36
D-Val
-
mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
5.9
D-Val
-
wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.7
glutaryl-7-aminocephalosporanic acid
recombinant mutant F54A, pH 7.5, 22°C
1.1
glutaryl-7-aminocephalosporanic acid
recombinant mutant F54S, pH 7.5, 22°C
1.5
glutaryl-7-aminocephalosporanic acid
recombinant wild-type enzyme, pH 7.5, 22°C
3.8
glutaryl-7-aminocephalosporanic acid
recombinant mutant F54Y, pH 7.5, 22°C
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
2.2
-
recombinant Strep-tagged wild-type enzyme, substrates are D-methionine and 6-dichloroindophenolate, pH 7.5, 37°C
2.6
-
recombinant Strep-tagged mutant R101A, substrates are D-methionine and 6-dichloroindophenolate, pH 7.5, 37°C
22.3
purified recombinant mutant F54S, substrate cephalosporin C, pH 7.5, 22°C
26.4
purified recombinant mutant F54A, substrate cephalosporin C, pH 7.5, 22°C
30.6
purified recombinant mutant F54Y, substrate cephalosporin C, pH 7.5, 22°C
8.5
purified recombinant wild-type enzyme, substrate cephalosporin C, pH 7.5, 22°C
additional information
additional information
intracellular activity of recombinant DAO in transformed Pichia pastoris, overview
additional information
-
intracellular activity of recombinant DAO in transformed Pichia pastoris, overview
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
7.5
8.5
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
6.5 - 10
-
pH 6.5: about 55% of maximal activity, pH 10.0: about 50% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
55
pI VALUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
physiological function
physiological role of D-amino acids and DAAOs, regulation of the nervous system, hormone secretion, and other processes by D-amino acids, detailed overview
physiological function
-
the enzyme is involved in the conversion of cephalosporin C
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). OXDA_TRIVR
356
0
39301
Swiss-Prot
Secretory Pathway (Reliability: 3)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
39000
40000
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
dimer
homodimer
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
crystallization of a single point mutant, X-ray diffraction structure determination and analysis at 1.8 A resolution
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
C106C108-(SO2H)
-
oxidatively modified enzyme shows 75% loss of activity
C108D
-
site-directed mutagenesis, in contrast to the wild-type enzyme, the mutant releases the cofactor in a quasi-irreversible manner and is therefore not stabilized by external FAD against loss of activity. Conformational properties and kinetics of C108D, overview
C108S
-
site-directed mutagenesis, in contrast to the wild-type enzyme, the mutant releases the cofactor in a quasi-irreversible manner and is therefore not stabilized by external FAD against loss of activity. Conformational properties and kinetics of C108S, overview
F258A
-
the improvement of catalytic efficiency with D-Tyr, D-Phe, and D-Leu for mutant enzyme F258A is 3.66, 11.7, and 1.5fold, respectively. The mutant is inactive with D-Ala, D-Ser, D-Lys, and D-Thr, while the activity with D-Tyr, D-Leu and especially with D-Phe significantly increases
F258S
-
the improvement of catalytic efficiency with D-Tyr, D-Phe, and D-Leu for mutant enzyme F258S is 1.7, 4.75, and 6.61fold, respectively. The mutant is inactive with D-Ala, D-Ser, D-Lys, and D-Thr, while the activity with D-Tyr, D-Leu and especially with D-Phe significantly increases
F258Y
-
the mutant is inactive with D-Val, D-Tyr, D-Ala, D-Ser, D-Lys, and D-Thr
F54Y
site-directed mutagenesis, the mutant shows 6fold improvement in kcat,app and about 2.5fold increase in Ki of glutaryl-7-aminocephalosporanic acid, the substitution improves the catalytic activity and thermostability of mutant DAAO compared to the wild-type enzyme. Heat treatment at 55° for 60 min does not decrease the activity of F54Y. The Tyr substitution might initiate hydrogen bond formation with the amino group of CPC and facilitate deamination
R110A
-
site-directed mutagenesis, the mutant shows activity similar to the wild-type enzyme, but shows increased temperature denaturation. Release of FAD is the dominant path of thermal denaturation of R110A
additional information
additional information
oxidation of Cys108 into cysteine sulfinic acid causes a global conformational response that affects the protein environment of the FAD cofactor. In comparison with the native enzyme, the mutation results in a fourfold-decreased specific activity, reflecting a catalytic efficiency for reduction of dioxygen lowered by about the same factor, and a markedly decreased propensity to aggregate under conditions of thermal denaturation
additional information
oxidation of Cys108 into cysteine sulfinic acid causes a global conformational response that affects the protein environment of the FAD cofactor. In comparison with the native enzyme, the mutation results in a fourfold-decreased specific activity, reflecting a catalytic efficiency for reduction of dioxygen lowered by about the same factor, and a markedly decreased propensity to aggregate under conditions of thermal denaturation
additional information
-
oxidation of Cys108 into cysteine sulfinic acid causes a global conformational response that affects the protein environment of the FAD cofactor. In comparison with the native enzyme, the mutation results in a fourfold-decreased specific activity, reflecting a catalytic efficiency for reduction of dioxygen lowered by about the same factor, and a markedly decreased propensity to aggregate under conditions of thermal denaturation
additional information
construction of point mutants with altered substrate specificity compared to the wild-type enzyme, the created mutant forms of TvDAAO are perfectly suitable for selective determination of D-Ser in excess of D-Ala, D-Asp, and D-Pro
additional information
oxidation of Cys108 into cysteine sulfinic acid causes a global conformational response that affects the protein environment of the FAD cofactor. In comparison with the native enzyme, the mutation results in a fourfold-decreased specific activity, reflecting a catalytic efficiency for reduction of dioxygen lowered by about the same factor, and a markedly decreased propensity to aggregate under conditions of thermal denaturation
additional information
oxidation of Cys108 into cysteine sulfinic acid causes a global conformational response that affects the protein environment of the FAD cofactor. In comparison with the native enzyme, the mutation results in a fourfold-decreased specific activity, reflecting a catalytic efficiency for reduction of dioxygen lowered by about the same factor, and a markedly decreased propensity to aggregate under conditions of thermal denaturation
additional information
-
oxidation of Cys108 into cysteine sulfinic acid causes a global conformational response that affects the protein environment of the FAD cofactor. In comparison with the native enzyme, the mutation results in a fourfold-decreased specific activity, reflecting a catalytic efficiency for reduction of dioxygen lowered by about the same factor, and a markedly decreased propensity to aggregate under conditions of thermal denaturation
additional information
-
immobilization of the recombinant enzyme on solid beads through affinity of its N-terminal Strep-tag to Strep-Tactin coated on insoluble particles, covalent attachment, re-usable in multiple cycles of substrate conversion, the surfactant Pluronic F-68 stabilizes DAO by protecting the enzyme from the deleterious effect of gas-liquid interfaces
additional information
development of a robust and highly active Pichia pastoris TvDAO whole-cell biocatalyst in a multistep engineering process for use in cephalosporin C conversion on industrial scale, overview. Usage of a fed-batch cultivation of the multicopy strain
additional information
-
development of a robust and highly active Pichia pastoris TvDAO whole-cell biocatalyst in a multistep engineering process for use in cephalosporin C conversion on industrial scale, overview. Usage of a fed-batch cultivation of the multicopy strain
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
6 - 8.2
stable from pH 6.0 to 8.2, above which a slight but continuous decrease in protein stability is observed
684631
4 - 11
-
at pH 4, about 40% of the maximal activity is observed for the immobilized DAO while no activity is detected for the soluble enzyme, at pH 11 immobilized DAO has 75% of the maximal activity, whereas the soluble DAOs retains only 13% of the maximal activity
685758
5 - 10
-
retains more than 80% of the maximal activity after incubation for 60 min at pH 5-10
685750
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
30 - 45
loss of secondary structure and inactivation occurs over a range of 30°C to 45°C, the melting temperature is at 41.4°C
40
stable up to 40°C, the enzyme has decreased stability at temperatures beyond and is completely inactivated at 65°C
45
fully stable up to 45°C (100% of residual activity after 30 min incubation)
25 - 50
-
added FAD does not stabilize DAO at and below 35°C, but it contributes up to 3.6fold extra stability to the enzyme activity at temperatures higher than 35°C, with a half-life of 60 h encapsulated DAO is 720fold more stable than the free enzyme under conditions of bubble aeration at 25°C, the soluble oxidase is stabilized by added FAD only at temperatures of 35°C or greater, at 50°C encapsulated preparations of the oxidase are much more stable than the free enzyme whose half-life is only 40 min
25 - 65
-
the activity profiles of soluble and immobilized DAOs at 25-65°C are similar, immobilized DAO exhibits a melting temperature of 60°C, which is 8°C higher than those for the soluble counterpart
30
-
increasing the temperature to 30°C does not affect the specific activity but enhances the volumetric enzyme activity by 22% due to its effect on cell growth
37 - 60
-
retains more than 80% of the maximal activity after incubation for 60 min at 37-60°C
40
42
-
50 h, pH 7.5, about 60% loss of activity of enzyme after multisubunit immobilization on highly activated glyoxyl agarose, protein concentration 0.067 mg/ml and 50 mM FAD, 95% loss of activity without FAD
50
55
additional information
-
thermal inactivation of D-amino acid oxidase from Trigonopsis variabilis occurs via three parallel paths of irreversible denaturation
40
-
50 h, pH 7.5, about 45% loss of activity of enzyme after multisubunit immobilization on highly activated glyoxyl agarose, protein concentration 0.2 mg/ml
50
-
half-life time of native enzyme is 1.69 h, half-life time of oxidatively modified enzyme C106C108-(SO2H) is 0.85 h. Thermal inactivation of D-amino acid oxidase from Trigonopsis variabilis occurs via three parallel paths of irreversible denaturation. One main path leading to inactivation is FAD release, a process whose net rate is 25-fold lower in the oxidized form of TvDAO. Cofactor dissociation is kinetically coupled to aggregation and can be blocked completely by the addition of free FAD. Aggregation is markedly attenuated in the less stable Cys108-SO2H-containing enzyme. A third, however, slow process is the conversion of the native enzyme into the oxidized form
50
-
inactivation of the enzyme at 50°C proceeds via two main pathways: partial loss of protein structure leading to a denatured holoenzyme, and reversible release of FAD cofactor generating inactive apoenzyme, mechanism, overview
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
His-tagged DAAO loses its flavin cofactor upon dilution or prolonged dialysis against FAD-free solvents leading to the catalytic inactivation
immobilisation on PEI 25 kDa Sepabeads and treatment with 0.5% glutaraldehyde solution at pH 7-8 and 25°C for 16 h leads to 160fold increased enzyme stability
carrier-free enzyme is entrapped in semipermeable microcapsules produced from the polycation poly(methylene-co-guanidine) in combination with CaCl2 and the polyanions alginate and cellulose sulfate, the effectiveness of the entrapped oxidase for O2-dependent conversion of D-methionine at 25°C is 75-95% of the free enzyme preparation
-
covalent modification of DAO using maleimide-activated PEG5000 yields a stable bionconjugate in which three exposed cysteine side chains are derivatized, the PEGylated enzyme shows approximately 3.3fold slowed dissociation of the FAD cofactor at 50°C, and this causes a 2fold thermostabilization of the enzyme activity
-
DAAO immobilized on a macro-porous carrier is higher in water-insoluble organic solvents than in water-soluble ones
-
enzyme immobilized on Amberzyme oxirane resin can be recycled more than 14 times without significant loss of activity
-
multisubunit immobilization on highly activated glyoxyl agarose marginally improves stability of the enzyme, 15-20fold, at a protein concentration of 0.0067 mg/ml. Dissociation of FAD is not prevented by immobilization
-
oriented immobilization of a chimeric oxidase maintains 80% of the original activity in microparticle-bound enzymes. Mildly permeabilized cells in which N-terminally tagged TvDAO in which a 12-amino-acid peptide is fused in frame to the N-terminus (TvDAOstrepN) is physically entrapped are employed for the batchwise conversion of D-Ala in an aerated enzyme reactor. The cells could be reused for at least three rounds of reaction with only small losses of activity. The free enzyme, however, is rapidly inactivated under the same reaction conditions (half-life: 1.2 h)
-
the catalytic efficiency of immobilized DAO (onto streptavidin-coated magnetic beads through the interaction between biotin and streptavidin) toward D-alanine is decreased by 56%
-
the surfactant Pluronic F-68 stabilizes immobilized DAO by protecting the enzyme from the deleterious effect of gas-liquid interfaces
-
OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
double DAO exhibits a half-life of about 30 min in the presence of 10 mM H2O2 which is 1.5fold longer compared to the native DAO
-
685750
in the presence of 10 mM H2O2, immobilized DAO exhibits a half-life of about 3 h giving 6fold greater stability than the soluble form
-
685758
oxidation of Cys108 into a stable cysteine sulfinic acid causes both decreased activity and stability of the enzyme
-
685045
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
enzyme intracellularly expressed under P(GAP) integrant in Pichia pastoris
-
Mono Q HR 10/10 column chromatography and Sephacryl S-200 gel filtration
-
recombinant N-terminally Strep-tagged enzyme to homogeneity from Escherichia coli strain BL21(DE3) by affinity chromatography
-
recombinant Strep-tagged wild-type and mutant enzymes from Escherichia coli strain BL21 (DE3) by ultracentrifugation, affinity and anion exchange chromatography
-
recombinant wild-type enzyme and mutants from Escherichia coli BL21(DE3)
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli strains BL21(DE3), TOP10F, JM105, JM109, and BL21
-
expression in Escherichia coli strain BL21(DE3), optimization of culture condition for the recombinant production of D-amino acid oxidase in Escherichia coli, overview
-
expression in Escherichia coli. TvDAO can be produced in Escherichia coli as a fully functional recombinant protein. They also show that microheterogeneity due to modification of Cys108 is avoided completely during production and purification
-
expression of His-tagged DAO (HDAO) in Pichia pastoris using the glyceraldehydes-3-phosphate dehydrogenase promoter. The maximal level of HDAO expression using the P(GAP) integrant is attained in 13 h and is equal to that obtained using the P(AOX1) integrant in 43 h.In-frame fusion of Saccharomyces cerevisiae alpha-factor secretion signal under a P(GAP) or P(AOX1) results in low-level secretion of active HDAO, which is not of practical use
-
expression of recombinant N-terminally Strep-tagged enzyme in Escherichia coli strain BL21(DE3)
-
expression of Strep-tagged wild-type and mutant enzymes in Escherichia coli strain BL21 (DE3), induction by DL-Ala
-
expression of wild-type enzyme and mutants F54Y in Escherichia coli BL21(DE3)
recombinant enzyme expression in Escherichia coli strain BL21(DE3), optimization of fermentation and cryopreservation of the recombinant Escherichia coli cells, detailed overview
-
stable expression in Pichia pastoris with 7fold enhancement of the intracellular level of oxidase activity, expression optimization by codon redesign as well as efficient subcellular targeting of the enzyme to peroxisomes
two cDNA genes were connected by a hexanucleotide linker and heterologously expressed in Escherichia coli to produce the corresponding double DAO with two subunits fused into a single polypeptide
-
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
biotechnology
the biotechnological applications of the enzyme range from biocatalysis to convert cephalosporin C into 7-amino cephalosporanic acid to gene therapy for tumor treatment
synthesis
the enzyme is used to develop biocatalysts for the production of 7-aminocephalosporanic acid from the natural antibiotic cephalosporin C, and to produce optically active L-methionine from D-amino acid with the yield of 100% in a cascade system of four enzymes, or to produce phenyl pyruvate from D-phenylalanine with the yield of 99%
biotechnology
synthesis
biotechnology
-
DAAO can be used to synthesize cephalosporin antibiotics
biotechnology
the enzyme is used as a biocatalyst in cephalosporin C conversion on industrial scale
synthesis
-
oxidation of cephalosporin C in the two-step formation of 7-aminocephalosporanic acid
synthesis
-
production of alpha-ketoadipinyl-7-aminocephalosporanic acid
synthesis
-
intracellular production of HDAO under P(GAP), followed by affinity purification and immobilization on oxirane resin, may serve as an effective process for the manufacture of immobilized DAO for industrial application
synthesis
-
immobilization of Trigonopsis variabilis D-amino acid oxidase on solid support is the key to a reasonably stable performance of this enzyme in the industrial process for the conversion of cephalosporin C as well as in other biocatalytic applications
synthesis
faster substrate turnover, reduced glutaryl-7-aminocephalosporanic acid inhibition and improved thermostability of the F54Y mutant compared to the wild-type enzyme render it a useful candidate in industrial production of semi-synthetic cephems
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Berg, C.P.; Rodden, F.A.
Purification of D-amino oxidase from Trigonopsis variabilis
Anal. Biochem.
71
214-222
1976
Trigonopsis variabilis
Szwajcer, E.; Mosbach, K.
Isolation and partial characterization of a D-amino acid oxidase active against cephalosporin. c from the yeast Trigonopsis variabilis
Biotechnol. Lett.
7
1-7
1985
Trigonopsis variabilis
-
Kubicek-Pranz, E.M.; Rhr, M.
D-amino acid oxidase from the yeast Trigonopsis variabilis
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7
104-113
1985
Trigonopsis variabilis
Schraeder, T.; Andreesen, J.R.
Properties and chemical modification of D-amino acid oxidase from Trigonopsis variabilis
Arch. Microbiol.
165
41-47
1996
Trigonopsis variabilis
-
Ju, S.S.; Lin, L.L.; Wang, W.C.; Hsu, W.H.
A conserved aspartate is essential for FAD binding and catalysis in the D-amino acid oxidase from Trigonopsis variabilis
FEBS Lett.
436
119-122
1998
Trigonopsis variabilis
Pollegioni, L.; Porrini, D.; Molla, G.; Pilone, M.S.
Redox potentials and their pH dependence of D-amino-acid oxidase of Rhodotorula gracilis and Trigonopsis variabilis
Eur. J. Biochem.
267
6624-6632
2000
Rhodotorula toruloides, Trigonopsis variabilis
Pollegioni, L.; Langkau, B.; Tischer, W.; Ghisla, S.; Pilone, M.S.
Kinetic mechanism of D-amino acid oxidases from Rhodotorula gracilis and Trigonopsis variabilis
J. Biol. Chem.
268
13850-13857
1993
Rhodotorula toruloides, Trigonopsis variabilis
Pilone, M.S.
D-Amino acid oxidase: new findings
Cell. Mol. Life Sci.
57
1732-1747
2000
Homo sapiens, Rhodotorula toruloides, Sus scrofa, Trigonopsis variabilis
Vikartovska-Welwardova, A.; Michalkova, E.; Gemeiner, P.; Welward, L.
Stabilization of D-amino-acid oxidase from Trigonopsis variabilis by manganese dioxide
Folia Microbiol. (Praha)
44
380-384
1999
Trigonopsis variabilis
Pollegioni, L.; Buto, S.; Tischer, W.; Ghisla, S.; Pilone, M.S.
Characterization of D-amino acid oxidase from Trigonopsis variabilis
Biochem. Mol. Biol. Int.
31
709-717
1993
Trigonopsis variabilis
Tishkov, V.I.; Khoronenkova, S.V.
D-Amino acid oxidase: structure, catalytic mechanism, and practical application
Biochemistry
70
40-54
2005
Candida parapsilosis, Fusarium oxysporum, Rhodotorula toruloides, Sus scrofa, Trigonopsis variabilis, Acrostalagmus luteoalbus, [Candida] boidinii (Q9HGY3)
Betancor, L.; Hidalgo, A.; Fernandez-Lorente, G.; Mateo, C.; Rodriguez, V.; Fuentes, M.; Lopez-Gallego, F.; Fernandez-Lafuente, R.; Guisan, J.M.
Use of physicochemical tools to determine the choice of optimal enzyme: stabilization of D-amino acid oxidase
Biotechnol. Prog.
19
784-788
2003
Rhodotorula toruloides, Trigonopsis variabilis
Pollegioni, L.; Caldinelli, L.; Molla, G.; Sacchi, S.; Pilone, M.S.
Catalytic properties of D-amino acid oxidase in cephalosporin C bioconversion: a comparison between proteins from different sources
Biotechnol. Prog.
20
467-473
2004
Rhodotorula toruloides, Sus scrofa, Trigonopsis variabilis
Verweij, J.; Vroom, E.D.
Industrial transformation of penicillins and cephalosporins
Rec. Trav. Chim. Pays-Bas
112
66-81
1993
Trigonopsis variabilis
-
Slavica, A.; Dib, I.; Nidetzky, B.
Single-site oxidation, cysteine 108 to cysteine sulfinic acid, in D-amino acid oxidase from Trigonopsis variabilis and its structural and functional consequences
Appl. Environ. Microbiol.
71
8061-8068
2005
Trigonopsis variabilis (Q6R4Q9), Trigonopsis variabilis (Q99042), Trigonopsis variabilis
Dib, I.; Stanzer, D.; Nidetzky, B.
Trigonopsis variabilis D-amino aacid oxidase: control of protein quality and opportunities for biocatalysis through production in Escherichia coli
Appl. Environ. Microbiol.
73
331-333
2007
Trigonopsis variabilis
Zheng, H.; Wang, X.; Chen, J.; Zhu, K.; Zhao, Y.; Yang, Y.; Yang, S.; Jiang, W.
Expression, purification, and immobilization of His-tagged D-amino acid oxidase of Trigonopsis variabilis in Pichia pastoris
Appl. Microbiol. Biotechnol.
70
683-689
2006
Trigonopsis variabilis
Dib, I.; Slavica, A.; Riethorst, W.; Nidetzky, B.
Thermal inactivation of D-amino acid oxidase from Trigonopsis variabilis occurs via three parallel paths of irreversible denaturation
Biotechnol. Bioeng.
94
645-654
2006
Trigonopsis variabilis
Lutz-Wahl, S.; Trost, E.M.; Wagner, B.; Manns, A.; Fischer, L.
Performance of D-amino acid oxidase in presence of ionic liquids
J. Biotechnol.
124
163-171
2006
Trigonopsis variabilis, Trigonopsis variabilis CBS 4095
Trampitsch, C.; Slavica, A.; Riethorst, W.; Nidetzky, B.
Reaction of Trigonopsis variabilis D-amino acid oxidase with 2,6-dichloroindophenol: kinetic characterization and development of an oxygen-independent assay of the enzyme activity
J. Mol. Catal. B
32
271-278
2005
Trigonopsis variabilis
-
Khoronenkova, S.V.; Tishkov, V.I.
High-throughput screening assay for D-amino acid oxidase
Anal. Biochem.
374
405-410
2008
Trigonopsis variabilis
Pollegioni, L.; Molla, G.; Sacchi, S.; Rosini, E.; Verga, R.; Pilone, M.S.
Properties and applications of microbial D-amino acid oxidases: current state and perspectives
Appl. Microbiol. Biotechnol.
78
1-16
2008
Fusarium solani (P24552), Rhodotorula toruloides (P80324), Rubrobacter xylanophilus (Q1AYM8), Glutamicibacter protophormiae (Q7X2D3), Trigonopsis variabilis (Q99042), [Candida] boidinii (Q9HGY3), Mycobacterium leprae (Q9RIA4)
Nidetzky, B.
Stability and stabilization of D-amino acid oxidase from the yeast Trigonopsis variabilis
Biochem. Soc. Trans.
35
1588-1592
2007
Trigonopsis variabilis
Arroyo, M.; Menendez, M.; Garcia, J.L.; Campillo, N.; Hormigo, D.; De la Mata, I.; Castillon, M.P.; Acebal, C.
The role of cofactor binding in tryptophan accessibility and conformational stability of His-tagged D-amino acid oxidase from Trigonopsis variabilis
Biochim. Biophys. Acta
1774
556-565
2007
Trigonopsis variabilis (Q99042), Trigonopsis variabilis
Nahalka, J.; Dib, I.; Nidetzky, B.
Encapsulation of Trigonopsis variabilis D-amino acid oxidase and fast comparison of the operational stabilities of free and immobilized preparations of the enzyme
Biotechnol. Bioeng.
99
251-260
2008
Trigonopsis variabilis
Kim, S.; Kim, N.; Shin, C.; Kim, C.
Optimization of culture condition for the production of D-amino acid oxidase in a recombinant Escherichia coli
Biotechnol. Bioprocess Eng.
13
144-149
2008
Trigonopsis variabilis, Trigonopsis variabilis CBS 4095
-
Wang, S.J.; Yu, C.Y.; Lee, C.K.; Chern, M.K.; Kuan, I.C.
Subunit fusion of two yeast D-amino acid oxidases enhances their thermostability and resistance to H2O2
Biotechnol. Lett.
30
1415-1422
2008
Rhodotorula toruloides, Trigonopsis variabilis
Wang, S.J.; Yu, C.Y.; Kuan, I.C.
Stabilization of native and double D-amino acid oxidases from Rhodosporidium toruloides and Trigonopsis variabilis by immobilization on streptavidin-coated magnetic beads
Biotechnol. Lett.
30
1973-1981
2008
Rhodotorula toruloides, Trigonopsis variabilis
Pollegioni, L.; Piubelli, L.; Sacchi, S.; Pilone, M.S.; Molla, G.
Physiological functions of D-amino acid oxidases: from yeast to humans
Cell. Mol. Life Sci.
64
1373-1394
2007
Chlorella vulgaris, Rattus norvegicus (O35078), Sus scrofa (P00371), Homo sapiens (P14920), Homo sapiens, Mus musculus (P18894), Rhodotorula toruloides (P80324), Cyprinus carpio (Q6TGN2), Trigonopsis variabilis (Q99042), [Candida] boidinii (Q9HGY3)
Pollegioni, L.; Sacchi, S.; Caldinelli, L.; Boselli, A.; Pilone, M.S.; Piubelli, L.; Molla, G.
Engineering the properties of D-amino acid oxidases by a rational and a directed evolution approach
Curr. Protein Pept. Sci.
8
600-618
2007
Sus scrofa (P00371), Sus scrofa, Homo sapiens (P14920), Homo sapiens, Rhodotorula toruloides (P80324), Trigonopsis variabilis (Q99042)
Liu, Y.; Li, Q.; Zhu, H.; Yang, J.
High soluble expression of D-amino acid oxidase in Escherichia coli regulated by a native promoter
Appl. Biochem. Biotechnol.
158
313-322
2008
Trigonopsis variabilis
Khoronenkova, S.V.; Tishkov, V.I.
D-amino acid oxidase: physiological role and applications
Biochemistry (Moscow)
73
1511-1518
2008
Homo sapiens, Sus scrofa, Rhodotorula toruloides (P80324), Trigonopsis variabilis (Q99042)
Kim, S.J.; Park, H.W.; Shin, C.H.; Kim, C.W.
Establishment of a cryopreservation method for the industrial use of D-amino acid oxidase-overexpressing Escherichia coli
Biosci. Biotechnol. Biochem.
73
299-303
2009
Trigonopsis variabilis, Trigonopsis variabilis CBS 4095
Dib, I.; Nidetzky, B.
The stabilizing effects of immobilization in D-amino acid oxidase from Trigonopsis variabilis
BMC Biotechnol.
8
72
2008
Trigonopsis variabilis
Mueller, M.; Kratzer, R.; Schiller, M.; Slavica, A.; Rechberger, G.; Kollroser, M.; Nidetzky, B.
The role of Cys108 in Trigonopsis variabilis D-amino acid oxidase examined through chemical oxidation studies and point mutations C108S and C108D
Biochim. Biophys. Acta
1804
1483-1491
2010
Trigonopsis variabilis, Trigonopsis variabilis ATCC 10679
Abad, S.; Nahalka, J.; Bergler, G.; Arnold, S.A.; Speight, R.; Fotheringham, I.; Nidetzky, B.; Glieder, A.
Stepwise engineering of a Pichia pastoris D-amino acid oxidase whole cell catalyst
Microb. Cell Fact.
9
24
2010
Trigonopsis variabilis (Q6R4Q9), Trigonopsis variabilis
Wong, K.S.; Fong, W.P.; Tsang, P.W.
A single Phe54Tyr substitution improves the catalytic activity and thermostability of Trigonopsis variabilis D-amino acid oxidase
New Biotechnol.
27
78-84
2010
Trigonopsis variabilis (Q6R4Q9), Trigonopsis variabilis
Komarova, N.V.; Golubev, I.V.; Khoronenkova, S.V.; Chubar, T.A.; Tishkov, V.I.
Engineering of substrate specificity of D-amino acid oxidase from the yeast Trigonopsis variabilis: directed mutagenesis of Phe258 residue
Biochemistry
77
1181-1189
2012
Trigonopsis variabilis
Kopf, J.; Hormigo, D.; Garcia, J.L.; Acebal, C.; de la Mata, I.; Arroyo, M.
Inhibition of recombinant d-amino acid oxidase from Trigonopsis variabilis by salts
Enzyme Res.
2011
158541
2011
Trigonopsis variabilis
Hou, J.; Liu, Y.; Li, Q.; Yang, J.
High activity expression of D-amino acid oxidase in Escherichia coli by the protein expression rate optimization
Protein Expr. Purif.
88
120-126
2013
Trigonopsis variabilis
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