Mammalian flavin-containing monooxygenase (FMO) as a source of hydrogen peroxide

Abstract

Flavin-containing monooxygenase (FMO) oxygenates drugs/xenobiotics containing a soft nucleophile through a C4a hydroperoxy-FAD intermediate. Human FMOs 1, 2 and 3, expressed in Sf9 insect microsomes, released 30–50% of O2 consumed as H2O2 upon addition of NADPH. Addition of substrate had little effect on H2O2 production. Two common FMO2 (the major isoform in the lung) genetic polymorphisms, S195L and N413K, were examined for generation of H2O2. FMO2 S195L exhibited higher “leakage”, producing much greater amounts of H2O2, than ancestral FMO2 (FMO2.1) or the N413K variant. S195L was distinct in that H2O2 generation was much higher in the absence of substrate. Addition of superoxide dismutase did not impact H2O2 release. Catalase did not reduce levels of H2O2 with either FMO2.1 or FMO3 but inhibited H2O2 generated by FMO2 allelic variants N413K and S195L. These data are consistent with FMO molecular models. S195L resides in the GxGxSG/A NADP+ binding motif, in which serine is highly conserved (76/89 known FMOs). We hypothesize that FMO, especially allelic variants such as FMO2 S195L, may enhance the toxicity of xenobiotics such as thioureas/thiocarbamides both by generation of sulfenic and sulfinic acid metabolites and enhanced release of reactive oxygen species (ROS) in the form of H2O2.

Characterization of a new Baeyer-Villiger monooxygenase and conversion to a solely N-or S-oxidizing enzyme by a single R292 mutation

“…Tynes et al [34] found hydrogen peroxide produced by the purified rabbit lung FMO (FMO2) at up to 41% of the total NADPH oxidized. More recently Siddens et al [35] published data on three human FMOs, where the percentage of molecular oxygen consumed that appeared as H 2 O 2 varied between 30 and 50% at a rate of 0.5-2.5 nmol/min/nmol FMO. However, all these uncoupling data are for mammalian FMO enzymes and to our knowledge no such data has been reported for the bacterial BVMO enzymes.…”

Flavin-containing monooxygenases in aging and disease: Emerging roles for ancient enzymes

“…1). Uncoupling has been observed in both the presence and the absence of substrate in vitro, resulting in "leakage" of hydrogen peroxide, and less frequently, superoxide (7). FMOs are notable for their "cocked and loaded" mechanism where they bind NAD(P)H and reduce FAD in the absence of substrate, creating an unusually stable C4a-hydroperoxyflavin ready to oxidize any substrate that accesses the active site (6).…”

“…The rate-limiting step of FMOs is thought to be the release of either H 2 O or NADP ϩ . The significance of FMOs' reaction kinetics, reactive oxygen species leakage, and effects on cellular NAD(P)H is not known (7,8).…”

Oxygen Metabolism in the Lung

“…The amine oxidases are members of the FAD monooxygenase (FMO) class, which are important sources of H 2 O 2 within the lung and play a critical role in xenobiotic metabolism (Francois et al, 2009;Siddens et al, 2014;Zhang et al, 2006). The FMOs of lung are distinctly different from liver, with FMO2 being the predominant form in adult lung; and there appears to be a distinct developmental aspect to their expression Hines, 2006).…”