A Key Role for Old Yellow Enzyme in the Metabolism of Drugs by Trypanosoma cruzi

Abstract

Trypanosoma cruzi is the etiological agent of Chagas' disease. So far, first choice anti-chagasic drugs in use have been shown to have undesirable side effects in addition to the emergence of parasite resistance and the lack of prospect for vaccine against T. cruzi infection. Thus, the isolation and characterization of molecules essential in parasite metabolism of the anti-chagasic drugs are fundamental for the development of new strategies for rational drug design and/or the improvement of the current chemotherapy. While searching for a prostaglandin (PG) F2α synthase homologue, we have identified a novel “old yellow enzyme” from T. cruzi (TcOYE), cloned its cDNA, and overexpressed the recombinant enzyme. Here, we show that TcOYE reduced 9,11-endoperoxide PGH2 to PGF2α as well as a variety of trypanocidal drugs. By electron spin resonance experiments, we found that TcOYE specifically catalyzed one-electron reduction of menadione and β-lapachone to semiquinone-free radicals with concomitant generation of superoxide radical anions, while catalyzing solely the two-electron reduction of nifurtimox and 4-nitroquinoline-N-oxide drugs without free radical production. Interestingly, immunoprecipitation experiments revealed that anti-TcOYE polyclonal antibody abolished major reductase activities of the lysates toward these drugs, identifying TcOYE as a key drug-metabolizing enzyme by which quinone drugs have their mechanism of action.

Ligand-induced Conformational Changes in the Capping Subdomain of a Bacterial Old Yellow Enzyme Homologue and Conserved Sequence Fingerprints Provide New Insights into Substrate Binding

“…Because SYE1 exhibits a preference for NADH, this theory cannot be extended to all OYE homologues. When we compare the utilization of reducing power among OYE homologues, we find that the ones preferring NADPH accepted both NADPH and NADH, but with a preference for the former (18,31,37,41,45). On the other hand, OYEs that use NADH cannot accept NADPH.…”

“…The OYE family has grown steadily in recent years and now features the bacterial nitro-ester reductases PETN reductase (8), glycerol trinitrate reductase (9), XenA/B reductase (10), the bacterial morphinone reductase (11), the YqjM from Bacillus subtilis (41), the plant oxophytodienoic acid reductases (12,13), several yeast OYEs (14 -17), and an enzyme involved in prostaglandin synthesis in Trypanosoma cruzi (18). Although these enzymes originate from different organisms and catalyze different reactions, they share several common functional characteristics.…”

Chagas disease: Present status of pathogenic mechanisms and chemotherapy

“…Many of the sequelae associated with T. cruzi infection are reminiscent of the effects of TXA2, including enhanced platelet adherence and aggregation in T cruzi-infected mice, where TXA2 levels are increased (Ashton et al, 2007;Cardoni and Antunez, 2004;Tanowitz et al, 1990). In T. cruzi, prostaglandin production, including TXA2 and PGF2 , is possible through a NADPH flavin oxidoreductase named "old yellow enzyme" (Kubata et al, 2002). T. cruzi derived TXA2 is important in modulating disease pathogenesis in the absence of host-derived TXA2.…”

Reduction of Quinones by NADH Catalyzed by Organoiridium Complexes

“…The simulated spectrum is characteristic of the menadione semiquinone radical anion (MC À ) with hyperfine coupling constants a H H5=H6=H7=H8 = 3.03 G, a H H3 = 0.65 G (Figure 2 b), which agree with a previous report. [11] The menadione radical anion formed here was stable for more than 20 h. Although some metal-quinone radical adducts have been reported, [5a] so far no superhyperfine splitting as a result of coupling with iridium nuclei was detected. Integration of the EPR signals (relative to a standard tempol (4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl) solution) showed that almost all the menadione (93 %) had been converted to menadione semiquinone.…”