Selenoxidation by flavin-containing monooxygenases as a novel pathway for beta-elimination of selenocysteine Se-conjugates

Previously, it was shown that beta -elimination of selenocysteine Se-conjug ates by rat renal cytosol leading to pyruvate formation was not solely cata lyzed by pyridoxal phosphate-dependent enzymes. It was hypothesized that se lenoxidation of the selenocysteine Se-conjugates, followed by syn-eliminati on, may be an alternative mechanism for pyruvate formation. In this study, selenoxidation of selenocysteine Se-conjugates was studied using rat liver microsomes and recombinant human oxidative enzymes. For all six selenocyste ine Se-conjugates that were tested, it was found that rat liver microsomal incubations led to the formation of pyruvate, whereas the corresponding sel enoxides were not observed. Microsomal pyruvate formation from Se-benzyl-L- selenocysteine (SeBC) was NADPH-dependent, but only marginally inhibited by several P450 inhibitors. Inhibition by methimazole and by heat pretreatmen t and stimulation by n-octylamine indicated that flavin-containing monooxyg enases are mainly responsible for pyruvate formation from the selenocystein e Se-conjugates in rat liver microsomes. In the case of S-benzyl-L-cysteine , the sulfur analogue of SeBC, pyruvate formation was not observed. For thi s substrate, a chemically stable sulfoxide could be observed, as previously described. By using recombinantly expressed human flavin-containing monoox ygenases and P450 enzymes, it was delineated that SeBC is selenoxidized by human FMOs, but not by human P450s. The k(cat)/K-m of selenoxidation was 3. 8-fold higher for FMO-1 than for FMO-3. In conclusion, selenoxidation of se lenocysteine Se-conjugates catalyzed by FMOs and subsequently syn-eliminati on has taken place as an alternative route for the formation of pyruvate fr om selenocysteine Se-conjugates. Although selenoxides are known to be easil y reduced by thiol compounds, microsomal pyruvate formation from SeBC was o nly 75% inhibited in the presence of an excess of glutathione. This indicat es that even in the presence of physiological concentrations of reducing th iol compounds, selenoxides of selenocysteine Se-conjugates may undergo syn- elimination to some extent. Whether selenoxides and/or selenenic acids that are formed are involved in the activity of chemopreventive selenocysteine Se-conjugates remains to be established.