How is the distal pocket of a heme protein optimized for binding of tryptophan?

Indoleamine 2,3-dioxygenase and tryptophan 2,3-dioxygenase catalyze the O(2) -dependent oxidation of l-tryptophan to N-formylkynurenine. Both are heme-containing enzymes, with a proximal histidine ligand, as found in the globins and peroxidases. From the structural information available so far, the distal heme pockets of these enzymes can contain a histidine residue (in tryptophan 2,3-dioxygenases), an arginine residue and numerous hydrophobic residues that line the pocket. We have examined the functional role of each of these residues in both human indoleamine 2,3-dioxygenase and human tryptophan 2,3-dioxygenase. We found that the distal histidine does not play an essential catalytic role, although substrate binding can be affected by removing the distal arginine and reducing the hydrophobic nature of the binding pocket. We collate the information obtained in the present study with that reported in the available literature to draw comparisons across the family and to provide a more coherent picture of how the heme pocket is optimized for tryptophan binding.

Oxidation of L-tryptophan in biology: a comparison between tryptophan 2,3-dioxygenase and indoleamine 2,3-dioxygenase.

The family of haem dioxygenases catalyse the initial oxidative cleavage of L-tryptophan to N-formylkynurenine, which is the first, rate-limiting, step in the L-kynurenine pathway. In the present paper, we discuss and compare structure and function across the family of haem dioxygenases by focusing on TDO (tryptophan 2,3-dioxygenase) and IDO (indoleamine 2,3-dioxygenase), including a review of recent structural information for both enzymes. The present paper describes how the recent development of recombinant expression systems has informed our more detailed understanding of the substrate binding, catalytic activity and mechanistic properties of these haem dioxygenases.

Reassessment of the reaction mechanism in the heme dioxygenases.

Indoleamine 2,3-dioxygenase (IDO) and tryptophan 2,3-dioxygenase (TDO) are heme enzymes that catalyze the O(2)-dependent oxidation of L-tryptophan to N-formyl-kynurenine. Previous proposals for the mechanism of this reaction have suggested that deprotonation of the indole NH group, either by an active-site base or by oxygen bound to the heme iron, as the initial step. In this work, we have examined the activity of 1-Me-L-Trp with three different heme dioxygenases and their site-directed variants. We find, in contrast to previous work, that 1-Me-L-Trp is a substrate for the heme dioxygenase enzymes. These observations suggest that deprotonation of the indole N(1) is not essential for catalysis, and an alternative reaction mechanism, based on the known chemistry of indoles, is presented.

A kinetic, spectroscopic, and redox study of human tryptophan 2,3-dioxygenase.

The family of heme dioxygenases, as exemplified by indoleamine 2,3-dioxygenase and tryptophan 2,3-dioxygenase, catalyzes the oxidative cleavage of L-tryptophan to N-formylkynurenine. Here, we describe a bacterial expression system for human tryptophan 2,3-dioxygenase (rhTDO) together with spectroscopic, kinetic, and redox analyses. We find unexpected differences between human tryptophan 2,3-dioxygenase and human indoleamine 2,3-dioxygenase [Chauhan et al. (2008) Biochemistry 47, 4761-4769 ]. Thus, in contrast to indoleamine 2,3-dioxygenase, the catalytic ferrous-oxy complex of rhTDO is not observed, nor does the enzyme discriminate against substrate binding to the ferric derivative. In addition, we show that the rhTDO is also catalytically active in the ferric form. These new findings illustrate that significant mechanistic differences exist across the heme dioxygenase family, and the data are discussed within this broader framework.