A Chemical Model of Tryptophan 2,3-Dioxygenase: Reactivity and Formation of a Reactive Intermediate in the Dioxygenation Processes.

Abstract

To understand the activation of molecular dioxygen by the heme-containing enzyme tryptophan 2, 3-dioxygenase (TDO), a chemical model was constructed, which permitted an identification of reactive intermediates through the use of ESR, optical spectra, cyclic voltammetry, and oxygenated product analysis. The temperature-dependent ESR measurement for the chemical model, which consisted of tetraphenylporphine iron(III) chloride (Fe(III)TPPCl), 3-methylindole (skatole; Sk), tetramethylammonium hydroxide (TMAOH), and molecular dioxygen, revealed the generation of two kinds of free radical species, as well as the formation of two ternary complexes in the ferric low-spin state. The radical species R1, with an ESR value of g=2.0045, was determined to be a Sk neutral radical derived from the substrate by MO calculation. The radical species R2, with an ESR value of g=2.015 was indicated to be a Sk-peroxide radical, consisting of a dioxygen molecule bound to the Sk neutral radical. A comparison of the ESR parameters of the two ternary complexes with those of previously reported complexes suggested that the present complexes are six-coordinate ferric low-spin species : one which is bound with two dioxygenated Sk anions at the heme-iron site ([A] : g1=2.236, g2=2.160, g3=1.965), and the other bound with a dioxygenated Sk anion and a methoxide anion ([B] : g1=2.311, g2=2.160, g3=1.954). These ternary complexes are assumed to be possible models for the transient hemoprotein-peroxide complexes. The formations of R1 and ternary complexes were indispensable for the dioxygenation of the substrate. On the basis of these results, a possible reaction mechanism is proposed.