Tryptophan 2,3-dioxygenase (TDO), an unrelated hepatic enzyme catalyzing the first step of tryptophan degradation, is expressed in many tumors and that this expression prevents tumor rejection by locally depleting tryptophan.

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TDO is a biologically significant enzyme. It catalyzes the first and committing step of L-Trp degradation in the kynurenine pathway. The kynurenine pathway constitutes the major route of de novo biosynthesis of NAD, which is one of the essential redox cofactors in all living systems. Excessive accumulation of many of the intermediate metabolites of this pathway can lead to numerous physiological and pathological conditions, including cataract formation, cerebral malaria, Alzheimer's disease, HIV infection, Huntington's disease and ischemic brain injury. TDO is responsible for oxidizing over 99% of the free L-Trp in intracellular and extracellular pools. Less than 1% Trp is metabolized via the serotonergic pathway, which synthesizes very important neurotransmitters such as serotonin and melatonin. Therefore, TDO plays a critical role in controlling the relative Trp metabolic flux between the serotonergic and kynureninic pathways. TDO was first discovered in mammalian liver by Kotake and Masayama in 1936 and was first characterized in rat liver at its purified protein level in 1955. A decade later, its isoform enzyme indoleamine 2,3-dioxygenase (IDO) was discovered and isolated from rabbit intestine.

IDO is primarily present in mammals, however and is ubiquitously distributed in various tissues of mammals except liver. TDO has a high specificity to L-tryptophan. It can also use a few tryptophan derivatives as substrate, for example, 6-fluoro-tryptophan. IDO has a much broader substrate specificity; it can utilize D-tryptophan, tryptamine and 5-hydroxytryptamine (serotonin) other than L-Trp as its substrate. The expression of IDO is induced by interferon-γ and linked to various immune-related pathophysiological conditions wherein TDO is induced by glucocorticoid hormones and regulated by its physiological substrate, L-Trp.

The cofactor of TDO/IDO

Hemoproteins perform a wide range of biological functions including oxygen transport and storage, electron transfer, monooxygenation, and reduction of dioxygen. However, they rarely express a dioxygenase activity as their native biological function. Dioxygenation reactions are typically catalyzed by non-heme metalloenzymes but TDO is the first described exception. As an oxygenation enzyme, TDO/IDO are distinctive members of the dioxygenase family as they utilize a histidine-ligated ferrous heme rather than a non-heme iron to carry out the oxygen activation and insertion reactions. The role of the metal in non-heme Fe-dependent dioxygenases is to facilitate a shift in electron density from the aromatic substrate to the bound oxygen. As for most of the non-heme iron dioxygenase, the non-heme iron active sites generally have two histidines, one carboxylate and two or three water ligands, which allow simultaneous binding of both substrate and dioxygen. The substrate and oxygen are activated and each presents some type of radical character so that the subsequent attack of oxygen on the aromatic substrate is spin allowed.

Most of the non-heme iron dioxygenases can be classified into extradiol and intradiol dioxygenase, which differ in their mode of ring cleavage and the oxidation state of the active-site metal. The ferrous extradiol enzymes activate O2 directly for reaction, whereas the ferric intradiol enzymes activate the substrate for O2 attack. Although they have different mechanisms, they share same common features, one of which is substrate directly binds to iron ion and forms the common enzyme-substrate complex. However, this mechanism cannot be applied to the enzymes with a heme cofactor because heme enzymes would not allow a simultaneous binding of the two substrates onto the Fe ion. In the ligand-bound crystal structure of TDO, the primary substrate (L-Trp) binds to a relatively hydrophobic pocket near (but not directly at) the Fe ion. The role of the Fe cofactor in the heme-dependent dioxygenase cannot be the same as the well-studied non-heme Fe counterpart. The dioxygenase activity of TDO must therefore proceed using a distinct new mechanism relative to that of the non-heme Fe-dependent dioxygenases. A recent discovery has exposed a third hemoprotein (PrnB) with dioxygenase activity. Therefore, a potential heme-dependent dioxygenase enzyme superfamily of which TDO is a prototype member has been suggested.