Biochemical characterization of a flavodiiron protein from bird parasite Histomonas meleagridis: superoxide as a reaction intermediate.

Histomonas meleagridis is a parasitic protozoan which causes histomoniasis (blackhead disease) in a wide range of birds, including domesticated chickens and turkeys, representing a significant health problem in avian veterinary medicine. Despite being classified as an anaerobic parasite, H. meleagridis can survive transient exposure to oxygen (O(2)) while little is known about the mechanisms that allow this organism to cope with exposure to varying O(2) levels. Inspecting the H. meleagridis genome revealed that this organism possesses two flavodiiron proteins (FDPs) belonging to classes A and F, which are known in other organisms to reduce O(2) to H(2)O. The class F FDP found in H. meleagridis (HmFDPF) has domain organization previously described for similar proteins and contains a middle rubredoxin-like and C-terminal NADH:rubredoxin oxidoreductase domains in addition to an N-terminal FDP core. Here, we present extensive biochemical characterization of this protein to improve understanding of the coordinated activities of multiple redox centers that enable NADH-dependent reduction of O(2) to H(2)O. We found that although superoxide is an HmFDPF reaction intermediate, only a very small amount of hydrogen peroxide (H(2)O(2)) is released as a byproduct. Moreover, HmFDPF does not possess NADH:H(2)O(2) oxidoreductase activity and the enzyme is prone to quick inactivation by H(2)O(2). In summary, our work expands our understanding of how microaerophilic protozoan parasites maintain their optimal redox state and minimize O(2) presence in their local environment, revealing mechanisms that can be exploited as drug targets against microaerophilic parasites.