Cloned (Comment) | Organism |
---|---|
recombinant expression of full-length His6-tagged hTDO protein in human liver HepG2 cell culture | Homo sapiens |
Crystallization (Comment) | Organism |
---|---|
purified enzyme in a ternary complex with the substrates L-Trp and O2 and in a binary complex with the product N-formylkynurenine, under-oil microbatch method under anaerobic conditions, mixing of 0.003 ml of 45 mg/ml protein in 50 mM Tris, pH 8.0, containing 150 mM NaCl and 10 mM L-Trp, and reduced with 2-fold molar excess of sodium dithionite, with 0.003-0.006 ml of precipitannt solution containing 50 mM sodium citrate, pH 5.6, 5% w/v PEG 3350, and 2% w/v Tacsimate, pH 5.0, X-ray diffraction structure determination and analysis at 2.5 A and 2.44 A resolution, respectively. Crystals of the Trp-bound binary complex are soaked in an O2-saturated precipitant solution supplemented with 20% v/v ethylene glycol at room temperature | Homo sapiens |
Protein Variants | Comment | Organism |
---|---|---|
Y175G | site-directed mutagenesis, the mutation leads to a 6fold slower multiple turnover velocity. In addition, pre-incubation of the Y175G mutant with 8 mM NFK retards the formation of the ternary complex by about 100fold and impedes Trp binding | Homo sapiens |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | stopped-flow kinetics | Homo sapiens |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Fe2+ | heme with Fe2+ coordinated by 4 N-atoms, overview | 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 | - |
- |
Reaction | Comment | Organism | Reaction ID |
---|---|---|---|
L-tryptophan + O2 = N-formyl-L-kynurenine | the dioxygenation reaction is initiated by a direct attack of O2 on the C2 atom of the L-Trp indole ring, catalytic mechanism, overview. Exo binding site for L-Trp, located about 42 A from the active site and formed by residues conserved among tryptophan-auxotrophic TDOs. Trp binding at this exo site does not affect enzyme catalysis but instead it retards the degradation of hTDO through the ubiquitin-dependent proteasomal pathway | Homo sapiens |
Source Tissue | Comment | Organism | Textmining |
---|---|---|---|
Hep-G2 cell | - |
Homo sapiens | - |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
L-tryptophan + O2 | - |
Homo sapiens | N-formyl-L-kynurenine | - |
? | |
additional information | substrate recognition and binding structures, overview. The EG segment in hTDO plays a critical role in promoting NFK release, thereby allowing Trp binding during multiple turnover | Homo sapiens | ? | - |
? |
Subunits | Comment | Organism |
---|---|---|
tetramer | - |
Homo sapiens |
Synonyms | Comment | Organism |
---|---|---|
TDO | - |
Homo sapiens |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
25 | - |
assay at | Homo sapiens |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
8 | - |
assay at | Homo sapiens |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
heme | with Fe2+ coordinated by 4 N-atoms, overview | Homo sapiens |
General Information | Comment | Organism |
---|---|---|
additional information | the exo site regulates hTDO cellular stability | Homo sapiens |