Cloned (Comment) | Organism |
---|---|
recombinant expression of N-terminally His6-tagged wild-type and mutant enzymes in Escherichia coli BL21(DE3) | Homo sapiens |
Crystallization (Comment) | Organism |
---|---|
purified full-length and truncated enzyme variants without heme cofactor, hanging drop vapor diffusion method, mixing of 0.001 ml of 20 mg/ml protein in 100 mM TrisHCl, pH 8.0, with 0.001 ml of reservoir solution containing 0.1 M sodium cacodylate, pH 6.0-6.5, 6-10% w/v MPEG 5000, and 10% v/v 2-propanol, addition of hexammine cobalt(III) chloride improves the diffraction significantly, 16°C, X-ray diffraction structure determination and analysis at 2.90 A resolution | Homo sapiens |
Protein Variants | Comment | Organism |
---|---|---|
F140A | site-directed mutagenesis, the mutant shows 0.25% of wild-type activity | Homo sapiens |
F72A | site-directed mutagenesis, inactive mutant | Homo sapiens |
H328A | site-directed mutagenesis, inactive mutant | Homo sapiens |
H76A, | site-directed mutagenesis, inactive mutant | Homo sapiens |
additional information | full-length hTDO is unstable, and various truncations are constructed. A truncation containing amino acids 19-388 is found to have good solubility and stability, the truncated mutant shows 80.45% of wild-type activity | Homo sapiens |
R144A | site-directed mutagenesis, the mutant shows 0.88% of wild-type activity | Homo sapiens |
S151A | site-directed mutagenesis, the mutant shows 9.08% of wild-type activity | Homo sapiens |
Y42A | site-directed mutagenesis, the mutant shows 0.5% of wild-type activity | Homo sapiens |
Y45A | site-directed mutagenesis, the mutant shows 1.13% of wild-type activity | Homo sapiens |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | Michaelis-Menten kinetics | Homo sapiens | |
0.135 | - |
L-tryptophan | pH 8.0, 25°C, recombinant wild-type enzyme | Homo sapiens | |
0.146 | - |
L-tryptophan | pH 8.0, 25°C, recombinant mutant S151A | Homo sapiens | |
0.153 | - |
L-tryptophan | pH 8.0, 25°C, recombinant truncated enzyme mutant | Homo sapiens | |
0.227 | - |
L-tryptophan | pH 8.0, 25°C, recombinant mutant Y42A | Homo sapiens | |
0.631 | - |
L-tryptophan | pH 8.0, 25°C, recombinant mutant R144A | Homo sapiens | |
0.661 | - |
L-tryptophan | pH 8.0, 25°C, recombinant mutant Y45A | Homo sapiens | |
1.531 | - |
L-tryptophan | pH 8.0, 25°C, recombinant mutant F140A | Homo sapiens |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
L-tryptophan + O2 | Homo sapiens | - |
N-formyl-L-kynurenine | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Homo sapiens | P48775 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant N-terminally His6-tagged wild-type and mutant enzymes from Escherichia coli BL21(DE3) by nickel affinity chromatography and gel filtration | Homo sapiens |
Source Tissue | Comment | Organism | Textmining |
---|---|---|---|
liver | mainly | Homo sapiens | - |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
L-tryptophan + O2 | - |
Homo sapiens | N-formyl-L-kynurenine | - |
? | |
L-tryptophan + O2 | cleavage of the C2-C3 bond in the indole moiety of L-Trp and incorporation of one oxygen molecule | Homo sapiens | N-formyl-L-kynurenine | - |
? | |
additional information | enzyme TDO is highly substrate-specific for L-Trp and the related derivatives 6-fluoro-Trp and 5-fluoro-Trp | Homo sapiens | ? | - |
? |
Subunits | Comment | Organism |
---|---|---|
dimer | apo hTDO | Homo sapiens |
More | hTDO must form an oligomer to exhibit activity. Active site structure, overview | Homo sapiens |
tetramer | holo-hTDO, dimer of dimers, in which the two C-shape dimers (AB and CD) are clamped perpendicularly to each other to form a tight tetramer | Homo sapiens |
Synonyms | Comment | Organism |
---|---|---|
hTDO | - |
Homo sapiens |
TDO | - |
Homo sapiens |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
25 | - |
assay at | Homo sapiens |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.01 | - |
L-tryptophan | pH 8.0, 25°C, recombinant mutant Y42A | Homo sapiens | |
0.032 | - |
L-tryptophan | pH 8.0, 25°C, recombinant mutant F140A | Homo sapiens | |
0.045 | - |
L-tryptophan | pH 8.0, 25°C, recombinant mutant R144A | Homo sapiens | |
0.062 | - |
L-tryptophan | pH 8.0, 25°C, recombinant mutant Y45A | Homo sapiens | |
0.105 | - |
L-tryptophan | pH 8.0, 25°C, recombinant mutant S151A | Homo sapiens | |
0.976 | - |
L-tryptophan | pH 8.0, 25°C, recombinant truncated enzyme mutant | Homo sapiens | |
1071 | - |
L-tryptophan | pH 8.0, 25°C, recombinant wild-type enzyme | Homo sapiens |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
8 | - |
assay at | Homo sapiens |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
heme | binding structure, overview | Homo sapiens |
General Information | Comment | Organism |
---|---|---|
additional information | eight residues play critical roles in L-tryptophan oxidation, i.e. Y42, Y45, F72, H76, F140, R144, S151, and H328. hTDO must form an oligomer to exhibit activity | Homo sapiens |
physiological function | tryptophan 2,3-dioxygenase (TDO) is one of the two key enzymes in the kynurenine pathway, it catalyzes the indole ring cleavage at the C2-C3 bond of L-tryptophan. This is a rate-limiting step in the regulation of tryptophan concentration in vivo. In addition to its role in protein synthesis, 95% of L-Trp in the human body is processed by the kynurenine pathway, leading to the production of nicotinamide adenine dinucleotide. Enzyme TDO is expressed in many tumor cells and is related to reduction of antitumor immune response | Homo sapiens |