120 - Indoleamine 2,3-Dioxygenase Is a Novel Mammalian Nitrite Reductase

Abstract

The heme enzyme indoleamine 2,3-dioxygenase 1 (IDO1) catalyzes the initial and rate limiting step of L-tryptophan (L-Trp) oxidation along the kynurenine pathway. IDO1 functions as a central immune regulatory enzyme with important implications for inflammation, infectious disease, allergic and autoimmune disorders, neuropathology and cancer. Here we identify IDO1 is a novel mammalian nitrite reductase that is capable of reducing nitrite to nitric oxide (NO) under hypoxia. Optical and Resonance Raman spectroscopy showed that incubation of dithionite-reduced, ferrous IDO1 protein (FeII-IDO) with nitrite under anaerobic conditions resulted in the time-dependent formation of a FeII-nitrosyl IDO1 species that was dependent on nitrite and IDO1 protein concentration. The bimolecular rate constant for IDO1 nitrite reductase activity was 5.1 M-1 s-1, which was comparable to that measured under identical conditions for myoglobin (3.7 M-1 s-1), an efficient and biologically important mammalian heme nitrite reductase. IDO nitrite reductase activity was also pH dependent but differed with myoglobin in that it showed a reduced proton dependency at pH>7. Electron paramagnetic resonance studies measuring NO production from nitrite showed that the conventional IDO1 dioxygenase reducing co-factors, ascorbate plus methylene blue, or ascorbate alone, supported IDO1’s nitrite reductase activity and the time-dependent release of NO in a manner modulated by substrate L-Trp and the IDO1 inhibitors 1-methyl-L-Trp and 1-methyl-D-Trp. These data identify IDO1 as a novel mammalian heme-based nitrite reductase that is capable of generating NO under hypoxia. IDO1’s nitrite reductase activity may have important implications for the immune regulatory actions of the enzyme when expressed within tumours and infected tissues that are characterized by hypoxia.