Comparative Studies of Human Indoleamine 2,3-dioxygenase and Tryptophan Dioxygenase

In contrast to the wide spectrum of cytochrome P450 monooxygenases, there are only two heme-based dioxygenases in humans, tryptophan dioxygenase (hTDO) and indoleamine 2,3-dioxygenase (hIDO). hTDO and hIDO catalyze the same oxidative ring cleavage reaction of L-tryptophan (L-Trp) to N-formyl kynurenine (NFK), the initial and rate-limiting step of the kynurenine pathway. Despite immense interest, the mechanism by which the two enzymes execute the dioxygenase reaction remains elusive. It was generally believed that the reaction proceeds via simultaneous incorporation of both atoms of dioxygen into the substrate. With resonance Raman spectroscopy, we have detected a key ferryl intermediate transiently populated during the dioxygenase reaction of hIDO. It supports a new mechanism, in which the two atoms of dioxygen are inserted into the substrate via a consecutive two-step reaction. Although this intermediate is not observable in the hTDO reaction, QM/MM studies indicate that both enzymes catalyze the reaction via the same mechanism, but the associated energy landscape is sensitive to the unique protein matrix surrounding the ligand and substrate in each enzyme. Comparative studies were conducted to reveal the substrate-ligand and ligand-protein interactions in the two important classes of heme-based dioxygenases.