Catalytic and Regulatory Mechanisms of Heme-based Dioxygenases

SUMMARY Human indoleamine 2,3-dioxygenases (IDO1 and IDO2) and tryptophan dioxygenase (TDO) are crucial hemeproteins responsible for catalyzing the first and rate-limiting step of the kynurenine pathway that serves as the primary metabolic route for breaking down L-tryptophan (Trp), the least abundant essential amino acid. By depleting Trp and facilitating the generation of bioactive kynurenine metabolites, these enzymes play a diverse role in immunomodulation. Accordingly, they have been implicated in a variety of pathological conditions, including cancer, and have been recognized as pivotal targets for therapeutic intervention. Extensive research in drug discovery targeting these enzymes has been initiated. The advances, however, have been significantly impeded by the poor understanding of the molecular properties of these enzymes. Our research program has made critical contributions in shaping our current understanding of these important dioxygenases at a molecular level, through unraveling the intricate details of the enzyme structures, substrate-selectivity, ligand-binding dynamics, dioxygenase activity and inhibitory mechanisms. We will continue our research endeavors in advancing the field by synergizing our efforts and expanding them to (i) define cellular metabolite- protein interactions and the associated regulatory mechanisms, (ii) unravel the catalytic diversity and the related enzymatic mechanisms, and (iii) delineate critical inhibitor-protein interactions for effective drug design. To achieve our goals, we will employ a multifaceted approach with a combination of spectroscopies, crystallography, calorimetry, high throughput mass spectrometry, protein engineering and computational simulations. The outcome of our research will (i) offer novel insights into the mechanisms by which the dioxygenases are regulated by cellular metabolites in the cellular milieu, (ii) fill in crucial knowledge gaps in structure-function relationships of the dioxygenases and heme oxygen chemistry, and (iii) expand our toolkit for rational design of enzyme-selective inhibitors.